Taxation Assignment Example | Topics and Well Written Essays - 1500 words

Taxation - Assignment Example Whether someone is employed or self employed depends upon the terms and conditions of relevant engagement. The tax and National Insurance contributions (NICs) rules do, however have special rules that apply to certain special categories of workers in certain circumstances. If you work for someone else it is important to know whether you are working for that person in employed capacity or in a self- employed capacity as an independent contractor.† (HMRC)ii Self employed taxpayers have the advantage of claiming wide range of expenses against the receipts of self employment services. â€Å"The decision to become self employed can also be affected by tax system.†(David G Blanch flower and Chris Shad forth)iii There is also an advantage of payment of income tax by instalments as against taxes being withheld by the employer under the PAYE scheme. It becomes extremely difficult in many cases to distinguish between self employment status and employee status of a tax payer. Whene ver any taxpayer is working on part time basis with more than one entity it is always the desire of HMRC to consider the tax payer under the employee status. This is because of easy and quick collection of taxes and NI under PAYE scheme on an income where only limited range of expenses can be claimed and thus providing higher amount of taxes. In fact these are the nature of activities of the taxpayer that decide the status of the tax payer. If there exists a contract of services then it is easy to consider taxpayer as an employee of the entity with whom the contract has been executed to render the services. But when such contracts exist for more than one entity the taxpayer would be treated as self employed rendering services to a number of entities. What will happen when there is no such contract? Under such conditions the issue is decided on basis of certain criteria. The main criteria are control over the performance of activities, fixidity of payment of remuneration even when th e entity is not earning, ownership of equipments used by the taxpayer during the performance of the duties, and exclusivity of providing services to one entity among other considerations. In our case Sam is now providing services exclusively to West London Company, and that is why HMRC wants tax him under the status of an employee and not under the status of as a self employed person. When you are not your own master and control of your activities and the performance is guided by the person providing you the remuneration, the status of the taxpayer is treated as an employee of the person providing him/ her the remuneration. â€Å"You are responsible for your own taxes and NI contributions† (Directgov)iv In the case of Sam his activities are now completely controlled by West London Company. Company is providing a fixed monthly remuneration of ?7500 even though he is operating from home. At the same time company is also providing him access to a desk, telephone, computer, and a lso stationery in the office. No reimbursement of expenses is allowed under the arrangement as the remuneration is fixed at ?7500 per month. There is exclusivity as Sam is also not providing services to any other entity or client. It should be noted that employment income is taxed on the basis of receipt of remuneration and not on accrual or earned basis which is applied when the taxpayer’

Sustainability in global business-Meeting the Challenge of Sustainable Essay

Sustainability in global business-Meeting the Challenge of Sustainable Business - Essay Example The human interactions that are pre-industrial with the natural system and the social system usually cause changes to the short-term and local scale. The present socio-ecological interactions are generating a wide spread in the global changes with changes that are temporary and far into the future. The harmful consequences that we face because of human forcing on the ecological and social systems may never find the solution required through single issues like solving the global poverty or resource efficiency (Blowfield, 2012, p. 18). The current processes and the societal structures are structured in a manner that they contribute systematically to the current directions that are unsustainable and causing the people to face the difficulty to inspire global unity around safeguarding the probabilities of the future options. The development of sustainability signifies the increase of the people’s room for maneuver. This requires an approach that is systematic in solving of problems and the strategic planning by identifying control for restoration in both ecological and human (Stoner and Wankel, 2010, p. 32). Many organizations are adopting policies and practices that are sustainable and they integrate them with the strategies in their organizations for the purpose of organization development (Schmidheiny, 1992, p. 214). Research has indicated that that most of the changes in the organizations efforts plateau after the initial period Research has of passion and was not able to become an integrated and sustained approach to the planning strategies and development. It is determined that changes that are unstained can be related to the current disclosure on the developments that are sustainable and focuses on the establishment of the evolving technology and instrumental policies such as what to do (Blowfield, 2012, p. 58). A complementary research

PERSONAL PERSPECTIVE for DRAMA OF DIVERSITY class, about these 2 Statement

PERSPECTIVE for DRAMA OF DIVERSITY class, about these 2 questions When did you first notice that you were different fr - Personal Statement Example I always felt that it was my moral obligation to help out others. I have probably given over 100 tutoring sessions in my lifetime free of charge. To me helping others was a truly gratifying experience. Back when I was 12 years old I would always play basketball during lunchtime and after school. I would practice alone for hours upon hours because I loved the game. My teachers always told me that I was a very special student. I have always loved reading about different subjects. When the internet became popular back in the 1990’s I became a more sophisticated student due to the fact that the internet provided me with an unlimited source of information. Since I was very young I was always very curious about learning new things. I believe that every human being has unique attributes and skills that can be used to make our society a better place. 2. A few years ago I was a member of an organization as a part-time worker. In this job one of my primary duties was giving refunds to t he employees that went on business trips. My supervisor was a very nice man that treated me with respect. After six months on the job my supervisor got promoted and he left the office to work at another division in another state. At that time the head of the office became my supervisor. From the start I did not like the way I was treated by this person. She though because I was young and inexperienced that I did not deserve any respect.

What are the potential benefits and pitfalls in promoting a new cinema Essay

What are the potential benefits and pitfalls in promoting a new cinema that places its emphasis on issues of identity and difference - Essay Example When motion pictures were first devised in the late 19th century, it was viewed as a medium for documentation of historical artefacts, in much the same way as still photographs were regarded. By the early 20th century the value of motion pictures as a form of entertainment and education was firmly established; however, it is only in the recent past, during the period of globalization, that the capability of film as a powerful tool for cultural dissemination has been recognized. Academic studies have turned to a determination of the effect of film in creating cross-cultural bridges between societies. A recent genre of film has been developed that explored the clashes among cultural influences and the ensuing resistance, assimilation, or adaptation of these elements. What emerges is the depiction of the evolution of identity, its definition in contemporary terms, and its differentiation from the status quo. There are both benefits and pitfalls to this new mode of filmmaking, which this paper will explore. It is however important to first describe the nuances of identity and differentiation, that becomes a defining force in the creation of motion pictures. Issues of identity and difference The driver for the depiction of cultural identity and its differentiation from â€Å"the other† is born by a universal need for public recognition of the distinctiveness of societies. The desire to preserve identity in one form or another is in response to the challenges of multiculturalism and the politics of recognition in democratic societies. The cultural context is valued by individuals as among their basic interests, and their right to this is universal. The problem resides, however, is in the reconciliation of the conflicting content of the various valued cultures. This implies limits that must be placed on political recognition of particular cultures (Amy Gutmann, 1994: 3). In Liberal democracies, citizenship cannot be regarded as a comprehensive universal identit y, for the reasons that: (1) people are unique, self-creating, and creative individuals; and (2) people are also â€Å"culture-bearing† whose cultures differ depending on their past and present identifications (Amy Gutmann, 1994: 7). Citizenship, therefore, is a unifying and equalizing factor in terms of rights; it does not, however, erase cultural distinctiveness. The liberal value of diversity should not be associated with the nullification, but rather the expansion and preservation of the â€Å"cultural, intellectual, and spiritual horizons of all individuals,† enriching and exposing us to differing intellectual and cultural perspectives, allowing for growth, exploration and enlightenment (Amy Gutmann, 1994: 9) The problem of cultural homogenization and cultural heterogenization is that there exists the indigenization of new cultural influences as they are constantly introduced into mainstream society (Arjun Appadurai). The new global cultural economy may no longer be understood in terms of simple models involving centers and peripheries, but instead as â€Å"a complex, overlapping, disjunctive order†. An elementary framework for exploring these disjunctures is suggested by Arjun Appadurai involves â€Å"five dimensions of global cultural flow†: (1) ethnoscapes; (2) mediascapes; (3) technoscapes; (4) finanscapes; and (5) ideoscapes. Each of these dimensions is a deeply imbued with historical, linguistic, and political context of nation-states, multinationals, diasporic communities (i.e., sharing common national or ethnic identity), and other such aggrupations. The five dimensions form the multiple worlds that are constituted by historically situated collective imaginations of persons and groups flung to various parts of the world (Appadurai, 2003). Deterritorialization generally is one of the major phenomena in contemporary times. It results from the mass movement of people of similar culture to other

Reducing Gases Emitted from Car Engines Essay Example for Free

Reducing Gases Emitted from Car Engines Essay The automobile industry has become one of the leading pollutants in most urban centers today. The emission of various gases from million of vehicles in roads has contributed to major global problem. Personal cars have emerged as the most notorious ones for this problem especially when they are moving at slow speeds or idling in traffic jams. This is the time when this man made gadgets produces the maximum emissions to the atmosphere with every body around that traffic absorbing the poison. Gasoline and diesel fuels combustion produces a mixture of hydrocarbons mainly carbon atoms, hydrogen as well as oxygen. The hydrocarbons are emitted as fragments of molecules of fuels which are burned partially. They do reactions with nitrogen oxides and in sunlight’s presence to create ground level ozone which becomes a main part of smog. This Ozone is harmful to human health as well as being toxic and sometimes causes cancer. Nitrogen Oxides (NOX) is produced in conditions of high pressure and temperature in the engine. Some nitrogen and oxygen atoms form nitrogen oxides after the reaction. NOX also contribute to creation of ozone as well as acidic rains which are harmful to the environment. NO2 which is much more potential than carbon dioxide as a green house gas is produced in the engine by catalytic conversion of heavy nitrogen gases in car exhausts systems. It contributes up about 7. 2 percent of gases responsible for global warming. Carbon monoxide, a colorless, odorless and poisonous gas is emitted from car engines after incomplete combustion of hydrocarbon-based fuels. It comprises of one atom of carbon and one oxygen atom interlinked together. Jeff Cohen asserts that, these results from having too low air-to fuel ratios in the car engine during the starting process of the car or when the car is not tuned on in the proper way. 1 This also takes place in higher altitudes in which case the amount of oxygen available for burning the fuel is greatly lowered. Transportation sources especially personal cars contributes to about two thirds of carbon monoxide. Carbon dioxide has been perceived as complete combustion product but of late it has been seen as a pollutant since it is a green house gas which traps the heat from the earth resulting to global warming. Of late, Bio-diesel has come up as one of the main renewable fuels and a provider of good opportunity for getting fossil fuel alternatives that assist in reducing emission of these green gases. It is capable of creating good blends in different ratios with diesel either fresh or the used up oil. This implies that Bio-diesel when used either completely or partially with diesel fuel in deferent vehicle engines brings about some economical, environmental as well as social merits. On the other hand, while Bio-diesel contributes to these changes, it also has some challenges on the uncontrolled substances which comprises of carbonyl compounds that are released in the process of emitting the combusted bio-diesels or bio-diesel and diesel blends. Since the fuels are quite different from the normal diesel fuels, they can change the injection as well as ignition processes. Due to this, the emission of green house gases such as CO, CO2, and NOX are greatly affected. This research aims at evaluating the effects of bio-diesel on combustion of diesel fuels as well as their emissions and control in order to obtain a picture of the opportunities and challenges provided by the bio-diesel fuels. 1. Jeff, C. Does Idling Your Car Hurt Gas mileage? Solve Your Problem. com, 2007, retrieved 28 May 2009, http://www. solveyourproblem. com/save-money-on-gas/idling-car-gas-mileage. shtml. Problem statement Most the current car engines are designed to use diesel fuel which is notorious for emitting green house gases. Other engines which employ diesel and bio-diesel mixture have not indicated the most appropriate ratio of mixing the fuels and the optimum speed and rack to emit the least amount of these gases to the atmosphere. The research therefore seeks to obtain the best ration of mixing diesel with bio-diesel or waste oil and the best rack to emit the least amount of gases to the atmosphere. Theses Statement Obtaining the most appropriate ratio of mixing fuels for car engines would assist in reducing the amount of green house emitted from these cars. This would help to aviate the global challenges imposed by the foresaid problem. Objectives Main Objective The main aim of the research is to reduce the amount of green house gases emitted from car diesel engines. Specific objectives These include obtaining the best ratio of mixing several fuels; either diesel with bio-diesel, diesel with waste oil or bio-diesel with waste oil or the best engine speed and rack for this. Scope of the research The research would investigate the most appropriate ratio of mixing different fuels but it has some limitations of financial resources and time. Literature Review There is much concern all over the globe to reduce the emission of green house gases by 10% in the near future. Much emphasis has been on emissions of these gases from transportation fuels. In 2005, the Indian Institute of Technology (IIT) New Delhi came up with a passenger car engine that operated on bio-diesel fuel. This was developed from modifications of the traditional Indica diesel engine that was made some few years ago. The engine used 10 per cent of bio-diesel fuel mixed with diesel to operate the engine. This development, however experiences some significant challenges in that it would take a lot of duration for any manufacture to come up with fully developed bio-diesel engine cars. In March 2009, United Environment and Energy LLC chemists came up with fixed-bed reactor design that was considered energy-efficient and high through in put continuous flow. It was perceived as cost-effective for production of algae oil bio-diesel. This was termed as the first economical approach in production of bio-diesel from algae oil. Although, the design has been able to reduce cost by 40% it did not reduce the emission of green house gases to an effective percentage. Tsinghua University researchers in China in May 2009 tried to use sugarcane juice as a good feedstock indented for production of bio-diesel if utilized to give support to alga Chlorella protothecoides through the process of heterotrophic fermentation. This produced oil content of about 53. 0% by weight of dry cell. In May 2009, Iowa State University Researchers presented a proposal on dissolution of waste polystyrene (PS) in bio-diesel. This aimed at making the bio-diesel for consumption in motor vehicles more so in diesel engines as an approach to recover energy from the waste plastic. The researchers pointed out that the use of polystyrene-bio-diesel blends would lead to increase in engine power thus reduced emissions of green house gases. This still intents increase polystyrene concentrations with 5% as per the study. To help reduce the emissions of green house gases, Tom Harkin and some US senators, in June 2009 proposed to the Environmental Protection Agency (EPA) to halt inclusion of in direct land use change effects as contributor to life-cycle greenhouse gas emissions in rule making indented for carrying out updated Renewable Fuels Standard that demands all bio-fuels to meet some defined life-cycle green house gas emission reduction targets prior to qualification. This research intents to come up with a more convenient approach of reducing the gases emitted from motor car diesel engines by obtaining the most optimum ratio of mixing bio-diesel, waste oil and diesel in car engine at the optimum engine speed. Research Methodology The research is conducted by experimenting the amount of green gases released from a diesel engine when diesel, bio-diesel, and waste oil are used at different to obtain different engine torques while the engine is increases from initial speed of 1500 rpm to 3200 rpm. The three fuels are used either purely or mixed with each other at different ratios while the rack is either maintained at 50% or at 100%. The amount of gases emitted for each condition is recorded in a table. The calves for the different gases emitted obtained at different speeds and racks are then plotted in several graphs. This is used to obtain the best fuel mixture and optimum speed and rack which release the least amount of gases to the atmosphere.

Mechanisms Of Conservative And Replicative Transposition Biology Essay

Mechanisms Of Conservative And Replicative Transposition Biology Essay Bacteriophage Mu is a temperate phage which adopts transposition pathway in its life cycle. Mu has the capability to integrate into numerous sites in host Escherichia coli genome and cause mutations due to its insertional activation. Mu transposes via two major pathways; conservative and replicative transposition though the molecular switch between the two mechanisms remain unknown. This review will focus on the comparisons between replicative and conservative transposition. The first part will discuss the similarities between the two mechanisms; donor DNA cleavage step and strand transfer step which involves nucleophilic attacks, generating single-strand nicks in Mu DNA and joining it to target DNA via one-step transesterification mechanism. The latter part will concentrate on the different characteristics in each transposition mechanism; in replicative transposition, the end product is duplication of transposon copy in both target and host DNA while in conservative transposition, a simple insertion of transposon is produced in the target DNA. 1. Characteristics of bacteriophage Mu Phage, derived from the Greek word phagein, literally means to eat. Bacteriophage Mu was named as such(find out who did) due its nature of infecting and inducing high levels of mutation in host bacteria Escherichia coli., hence the name Mu for mutator. The dual nature of Mu transposon and virus has made it as the archetypal model of studying phage genetics. Bacteriophage Mu is a temperate phage of E. coli which employs the transposition mechanism in its life cycle. Transposition can either be conservative (excising the transposon and inserting it into bacterial chromosome) or replicative (transposon copies are produced in both transposon and bacterial chromosome). Both mechanisms will be discussed extensively later in this article. Unlike the phage ÃŽÂ », insertion of Mu genome into the target site proceeds in a randomly manner which makes it an excellent mutator. Fig. 1: The life cycle of bacteriophage Mu(5). The life cycle of phage Mu is shown schematically in Fig. 1 above. Bacteriophage Mu infect susceptible host cell by adsorption and then, injects its linear viral genome. Once inside the host cell, the linear genome does not circularized(4,5,19), unlike in phage ÃŽÂ ». In either case of lytic or lysogenic phase, Mu integrates its DNA into the host genome via conservative transposition(16,19). This is observed differently in phage ÃŽÂ » where the infecting phage DNA will be integrated into host genome only during lysogenization(19). An enzyme called transposase, encoded by MuA gene in the phage genome, is absolutely crucial to carry out this conservative transposition step. Phage DNA is inserted at multiple sites in a bacterial genome which lead to the assumption that the insertion occur by a random manner(8). However, there are several factors that influence target site selection such as MuA protein efficiency and transposition immunity(15). After integration, Mu usually adopts a quiescent prophage lifestyle(lysogenic phase). The preference between lysogenic and lytic phase in Mu life cycle is dependent on its stability in the lysogen and lysogenic repressors. However, lysogens of Mu phage sometimes enter the lytic phase though this is a rare event. When induced, usually by using temperature-sensitive repressor mutants of phage Mu and subject it at 42ËÅ ¡C, the lysogen will enter lytic cycle. When the lysogenic repressor is inactivated, Mu transposes via replicative transposition, producing copies of phage genome which will be packaged into new virions. The virions then lyse the host cell and infect new hosts. Bacteriophage Mu virions comprised of icosahedral head(diameter 54nm), a baseplate, a contractile tail and six short tail fibres(5). Fig. 2: Simplified cartoon illustrating packaging of Mu genome. Typical length of phage Mu DNA is approximately 37kb long. Additional 2 kb of host DNA is incorporated during DNA packaging which is shown as flanking each end of the integrated Mu genome, with most of it at the right end. Unique sequences of host DNA and at the right end of the packaged DNA is dependent on initiation site of packaging in the host DNA(24). Fig. 3: Physical and genetic map of bacteriophage Mu. Solid black lines represent Mu DNA while the boxes at the two ends indicate flanking host DNA sequences. Mu genes (indicated in block letters) and their corresponding translational products are as indicated(19). A typical size of wild-type phage Mu DNA is about 37.5 kb, however each phage capsid can accommodate up to 39 kb long. Phage genome has a pac site which serves as the starting point in packaging of the phage DNA, located within attL(5). The initiation cleavage by phage enzyme terminase occurs upstream of the phage pac site, which includes host sequence of about 50-150bp flanking the left end. Second cleavage initiated when a complete filling of capsid is achieved, which includes 0.5 kb to 2 kb of host sequence flanking the right end(1). Genetic and physical map of phage Mu is illustrated in Fig. 3. Bacteriophage Mu utilizes headful mechanism strategy, which confer variable lengths of host DNA flanking the left ends of Mu DNA depending on the initiation site of genome packaging(Fig. 2). 2. Transposition mechanism (E) (D) (C) (B) (A) Fig. 4: Modes of bacteriophage Mu transposition. (A), (B) and (C) are the common steps in both conservative and replicative transposition of phage Mu. In conservative and replicative transposition, phage Mu will follow-up step (D) and (E) respectively. Curved arrows indicate nucleophile attack, transferring the 3-OH ends to the staggered 5-phosphate ends of target DNA. Dentate lines (XXXX) indicate target DNA sequences which are duplicated during transposition (16). Numerous in vitro studies have been conducted to study the mechanism of transposition, and usually mini-Mu elements are used. A minimal Mu element consists of a selectable gene, a plasmid replication origin and essential Mu ends(2). The mechanism of transposition is discussed in respect to an in vitro system from this point onwards unless stated otherwise. Following discussion on transposition mechanism are based on Shapiro model(22) as it has been widely accepted as the golden model in this field. The current known modes of transposition is divided into two: non-replicative (conservative) and replicative transposition. Both strategies utilize the same mechanism up to point (Fig. 4C) where each strategy employs different mechanism, producing different end products. A simple insertion of transposon is generated in target DNA by conservative transposition (Fig. 4D) while two copies of transposon formed in both donor and target DNA by replicative transposition (Fig. 4E). Point A to C are considered as the similar features in both conservative and replicative transposition while point D and E is the distinction between the two modes of transposition. Therefore, mechanisms involved in point A,B and C are discussed in context of both replicative and conservative transposition, which comprises of DNA cleavage step and strand transfer step. Sequential stages of both cleavage and strand transfer steps are illustrated in Fig. 4. 2.1 Donor DNA cleavage step Two critical chemical steps in both transposition pathways are donor DNA cleavage step and DNA strand transfer step(5,8). The donor DNA cleavage step is initiated when water molecules within an active site act as nucleophiles, and attack phosphodiester bond in DNA backbone at each of the transposon end(4,5). The cleavage step involves a direct hydrolysis of phosphodiester bond by water, and not by covalent enzyme-DNA intermediate(17). The phosphodiester bond is cleaved at the flanking host-transposon DNA boundary. 3-hydroxyl (OH) ends of the Mu DNA are exposed at the end of the cleavage step. Strand transfer results in fusion of target and donor DNA, which forms an intermediate molecule (8). The process (simplified in Fig. 4C) follows the Shapiro model(22). Bacteriophage-encoded proteins, MuA protein (transposase) and MuB protein (ATPase) are required for transposition. Other requirements to ensure efficiency of transposition are accessory proteins such as host-encoded DNA bending proteins called hydroxyurea (HU) and integration host factor (IHF)(8). The inverted repeats at the end of donor DNA, and target sequence on bacterial chromosome are also important in transposition mechanism. The assembly of higher order protein-DNA complexes called transposome has been identified by in vitro studies(6). A three-site synaptic complex called the LER complex comprising right and left ends of Mu and transpositional enhancer, was formed in the beginning of transposition in vitro(23). MuA protein binds to MuA binding site at the ends of Mu DNA as monomer, and subsequently function as tetramer of MuA (transposase). Host IHF and HU protein were found to aid in formation and stabilisation of LER complex. The LER complex is relatively unstable and so, is rapidly converted into stable synaptic complex (SSC), also known as type 0 complex(17). This is the critical checkpoint before any chemical reaction is carried out as it is the rate-limiting step of cleavage reaction(6). A stable synapse between tetramer of MuA and the two ends of Mu DNA is made but no cleavage is initiated yet at this point. Nonetheless, the active site is structurally occupied to the region around the scissile phosphate while the flanking DNA are destabilized upon formation of the SSC complex(6). In addition to formation of a stable synapse, the Mu ends needs to be properly-oriented, a super coiled DNA topology, and accessory DNA sites are also important to proceed to the next step. Formation of SSC usually is short-lived in presence of Mg2+ but can be accumulated in presence of suitable divalent cations such as Ca2+,which promotes the formation of SSC(8,17). Next, SSC is converted into a type 1 transposome complex, also called as cleaved donor complex(CDC)(9). The 3 ends of Mu DNA are nicked in presence of Mg2+. Two subunits of MuA tetramer, that are associated with the sites that undergo cleavage, assemble in trans arrangement which favours the strand transfer reaction(5). The formation of CDC can then be thought as the result of donor DNA cleavage step. Type 1 transposome complex exhibits greater stability than the type 0 complex though MuA forms structural and functional core in both transposome complexes(6). In addition of stably bound tetramer of MuA proteins, there are loosely associated MuA proteins present in the CDC as well. In absence of MuB protein, MuA tetramer is unable to promote strand transfer reaction unless these extra MuA proteins are present. MuB protein is an ATP-dependent DNA-binding protein, which also acts as an allosteric activator of Mu transposase (MuA proteins)(21). Transposition can still proceed in absence o f MuB proteins, but MuA protein by itself is only 1% efficient(3). 2.2 Strand transfer step A hallmark of this step is the formation of strand transfer complex (STC), also known as type 2 transposome complex. The end product of STC is formation of a branched molecule(Shapiro intermediate) which is characterized by a covalent interaction between donor DNA and target DNA via 5bp single-stranded gaps and its ÃŽÂ ¸ structure(22). MuB protein first captures a target molecule and bring it to the vicinity of the transposome complex, forming a TC complex(6). Formation of TC complexes rapidly undergo one-step transesterification reaction, which is the rate-limiting step in the strand transfer step. Interestingly, recruiting of target molecules by MuB proteins and formation of TC complexes can occur at several time point during the reaction pathway(6). This is a particularly efficient step to maximize transposition potential as it would speed up rate of strand transfers during transposition. The free 3-OH ends produced from the cleavage step act as nucleophile and attack phosphates of target DNA at the 5 ends. 5-nucleotides long offset nicks are made in the target DNA, generating a staggered arrangement(3). At this stage, the MuA proteins(transposase) are still tightly bound to the branched molecule with single stranded gaps. This pose an obstruction for the assembly of replication fork by host replication factors. The structure of the branched molecule is simplified in (C) of Fig. 4. The forming of this intermediate molecule serves as the critical point which distinguish between conservative and replicative transposition. A widely accepted model is that the resolving of this co-integrate molecule by a special resolvase complex leads to double copies of transposon being made in both donor and target site(REFerence). This is by definition, a replicative transposition pathway. Thus, the strand transfer complex is destabilized and disassembled by a system of eight E. coli host molecular proteins (DnaB helicase, DnaC protein, DnaG primase, DNA polymerase II, single-strand binding protein, DNA gyrase, DNA polymerase I and DNA ligase) and molecular chaperon called ClpX, producing cointegrates(13). This transition from transposome complex to a replisome results in duplication of 5-bp target DNA sequences flanking both ends of Mu DNA. Alternatively, if the bacteriophage Mu is to enter the conservative pathway, the co-integrate molecule is repaired or processed without performing Mu DNA replication. The end product of STC in a conservative transposition is a simple insertion of single mini-Mu element inserted into the target DNA(8). However, the mechanism of this model is poorly understood. Fig. 5: Transposome complexes involved during DNA cleavage complex and DNA strand transfer. (A) A plasmid (gray line) bearing donor mini-Mu element (black line) DNA in the in vitro system is negatively coiled. (B) In presence of host HU protein, Mu A protein bind to the two ends of Mu DNA forming a stable synaptic complex (not shown). Assembly of MuA tetramer produces a nick at each ends of Mu DNA, creating a cleaved donor complex (CDC). (C) Nicked 3 ends of Mu DNA are joined together to target DNA in presence of MuB protein forming a strand transfer complex (STC). MuA tetramer is still tightly bound to the Mu ends in the STC. (D) In replicative transposition, a cointegrate molecule is produced when replication of target DNA initiated from the 3 Mu ends by host replication machinery (13). 3. Replicative transposition Replicative transposition was first suggested by Ljungquist and Bukhari (1977) to occur in situ after induction of lysogens, which means that the Mu prophage was not excised from host chromosome during transposition(14). The lysogens were digested with restriction enzymes which cleaves both host and Mu DNA at specific restriction sites. Two of the fragments from the restriction digests contain both host and Mu DNA, which corresponds to junctions between host and prophage DNA, suggesting that prophage DNA is replicated in situ of host chromosome(19). Several genetic and biochemical predictions made in the Shapiro model have been demonstrated in both in vivo and in vitro studies, hence this model is accepted as a plausible mechanism to explain transposition in phage Mu. Numerous techniques have been done to study the direction of replication of Mu DNA during transposition. Results obtained by annealing of Okazaki fragments to separated strands of Mu DNA shows that more than 80% of Mu molecules replication proceed from left to right end(11,19). Electron microscopical observation of mini-Mu element shows that replicating molecules in vitro replicate from both ends in equal probability'(11,19). Replication of Mu DNA is accepted to be predominantly unidirectional, that is from left towards the right end(20). Intramolecular replication pathway can result in inversion, deletion, and simple insertion while intermolecular events can produce co-integrate molecules(19). In the case of Mu transposition, formation of co-integrate molecule needs to be resolved in order to produce two replicons; one molecule contains transposon and target DNA while another molecule contains transposon and donor DNA(10). 4. Conservative transposition The main characteristic of conservative transposition is that phage DNA is not replicated prior to integration. Upon infection of a susceptible host cell (usually E. coli), Mu employs conservative, or also called non-replicative transposition to transfer its genome to the target site. As discussed earlier, conservative transposition pathway follows single strand nicks at the 3 ends of Mu DNA, of which the exposed 3-OH ends join to the staggered cut target DNA at the 5ends forming a co-integrate molecule. The co-integrate or so-called Shapiro intermediate is repaired and generates a simple insertion in the target DNA though the mechanism is still poorly understood. Shapiro model emphasized on single-stranded nicks at Mu ends, joining of Mu to a staggered double-strand break in target DNA, formation of an intermediate molecule, and shedding of heterogeneous of previous host DNA sequences after ligation in conservative pathway(22). On the other hand, Morisato and Kleckner (1984) proposed a different mechanism based on results with Tn10 transposition. Their model is double-stranded cleavages at the transposon ends generating an excised transposon, which then circularizes via ligation on one of the strands(18). It predicts shedding of host sequences from the Mu DNA ends before ligation into the new target DNA. Study of Mu transposition using plasmid substrates in vitro produced results in favour of the Shapiro model, and hence this model has been widely accepted and used in studies. Fig. 6: A model of conservative transposition which utilizes double-strand cleavages during integration. (A) Transposase bind to the inverted repeats at Mu-host boundary sites and cleaves off the transposon away. (B) Transposase made a staggered cut at target sequence of which exposed 3-OH ends of transposon attacks 5-phosphate ends of the host (not shown). The transposon then joins to the host sequence. Duplicated target sequence of 5-bp are completed by host replication machinery (7). The debate on single-strand or double-strand cleavage however does not end there. If phage Mu were to utilize the Shapiro model of transposition during integration (the well-established cointegrate mechanism), the flanking host sequences would remain bound to Mu ends. This would clearly pose a problem as subsequent target-primed replication of the linear integrant would not work, or simply break the chromosome(1). Evidently, results from in vitro experiments are against this as the transposition end products contain transposon, suggesting a complete transposition process have been accomplished. So, does the infecting Mu DNA utilize the Shapiro model where the cointegrate molecule gets processed and repaired, prior to replication at the flanking sequence? Or does it follow a cut-and-paste mechanism where both strands of Mu DNA gets cleaved off from the flanking host DNA sequence (as illustrated in Fig. 6), where no cointegrate molecule is generated, which eventually means, there is no need for resolve by replication? An in vitro experiment was done by Au et al. (2006) to observe the fate of flanking host DNA sequences upon phage Mu infection. Specific markers specific to the infecting phage Mu DNA as well as the donor host (lacZ/proB) were used. These markers were acquired from the host in which the phage had been propagated but absent in the host being infected(1). Upon infection of plasmids by bacteriophage Mu, signal for flanking sequences and Mu DNA were detected in the chromosome at the same time point (approximately at minute 8), which correspond to the integration time point of Mu. Subsequent expression of lacZ and proB were detected maximally at minute 15, significantly reduced at minute 30 and by minute 50, expression were halted(1). Maximal expression at minute 15 most likely corresponds to climax of integration of the infecting phage population. These findings strongly suggest that flanking sequences get integrated together with Mu DNA into the new target site and are subsequently, rem oved by a special mechanism(which explained the undetectable expression at minute 50). This then proves that infecting phage Mu employs an alternate cointegrate mechanism (also called as nick-join-process mechanism) in conservative transposition pathway, where the Mu DNA undergo single-strand nicks, joins to the target DNA, and repaired before replication of the 5-bp gap left by the flanking sequence(1). The mechanism of removal and repair of host flanking sequence however, remains ambiguous. Conclusion Dual nature of bacteriophage Mu, a transposable element and a virus, is certainly interesting but what is more fascinating is that it utilizes both replicative and non-replicative transposition throughout its life cycle. The former mechanism produces a transposon copy in both donor and target DNA while the latter usually generates a simple insertion of transposon in the target DNA, leaving a gap in the host DNA which most likely will get degraded. In the early stages, both replicative and conservative transposition pathway share a similar mechanism. Regardless of the transposition pathway, infecting Mu DNA during the first round of infection will integrate its DNA into the target chromosome via two critical steps; donor DNA cleavage step and strand transfer step. Mu uses a phosphoryl transfer involving nucleophilic attacks of water on phosphodiester bonds of Mu DNA, producing single-strand nicks. A second nucleophilic attack by exposed 3-ends of Mu DNA on 5-ends of target phosphodiester bonds, which then joins the Mu DNA to target DNA via one-step transesterification mechanism. A series of transposome complexes are formed throughout these processes including Mu-encoded MuA proteins(transposase) and MuB proteins(ATPase). A cointegrate is produced in both pathways but in replicative transposition, this intermediate molecule is resolved producing two replicons with transposon copy in each molecule. In conservative transposition, the cointegrate is repaired generating a simple insertion in the target DNA. Hence, it is more accurate to name conservative transposition as nick-join-process rather than the conventional cut-and-paste mechanism as the latter suggest double-strand nicks at the transposon end, which has been proven inaccurate by in vitro experiments. Both transposition pathways have been compared extensively in this review but much of functional core of the mechanisms remain to be understood. (2944 words)

Health Care Essay -- Papers Insurance Work Essays

Health Care Abstract "Health Care", This is becoming more important in today's world of corporate streamlining, downsizing, and increased health care costs. A compensation package for the employees of today is more valuable than it has ever been. The average employee no longer is just concerned with the wages or salary they receive, their attention has been drawn to the overall compensation package. As health care costs rise the importance of an enrollment in a health care plan grows. Health care will continue to be an important factor for employees as long as costs continue to grow. I. Health Care. Starting from the time a person leaves their parents and as they enter the workforce and continue through life healthcare is an issue. Security is a basic human need and health care is one of the biggest parts of that security. It is important to know if one becomes ill or injured that treatment is readily available to them. This includes not only the employee but also the employee's family and or life partner. Health care here in the United States is not socialized and therefore is provided through the purchase of health care plans entirely by the employer or with co-payments from the employees. A. Providing health care programs is not always a guaranteed part of a compensation package when employed. Depending on the organization and its total compensation package health care may or may not be included. The organization may chose to provide the health care to the employees free of charge or they may be required to contribute a significant amount each month. 1. An employer designs compensation packages to attract qualified employees who have the knowl... ...f the organization a viable health care plan is necessary. To ensure plans are within an acceptable expense it is necessary to educate employees in preventive health care measures. Healthy employees make for less expenditures of plans and more productivity. Reviewing the facts and reports it is clear that health plans are a necessity rather than a nicety to help the organization meet its goals. References: Compensation Management in a Knowledge-Based World, (2000). By Richard I. Henderson. Henry Holt and Company Inc., (1999). "Forbidden Love" by Gary B. Nash http://www.ebri.org/ http://www.ifeb.org/icnewind.html http://www.nbch.org/ http://www.healthgrades.com http://www.presbyterian.org/employers.html http://www.healthcareresource.net/healthplan.html http://www.ftp.bls.gov/publ/news.release/hce.txt