Isozyme

Isozymes (also known as isoenzymes or more generally as multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters (e.g. different KM values), or different regulatory properties.The existence of isozymes permits the fine-tuning of metabolism to meet the particular needs of a given tissue or developmental stage.In biochemistry, isozymes (or isoenzymes) are isoforms (closely related variants) of enzymes. In many cases, they are coded for by homologous genes that have diverged over time. Although, strictly speaking, allozymes represent enzymes from different alleles of the same gene, and isozymes represent enzymes from different genes that process or catalyse the same reaction, the two words are usually used interchangeably. Isozymes (also known as isoenzymes or more generally as multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters (e.g. different KM values), or different regulatory properties.The existence of isozymes permits the fine-tuning of metabolism to meet the particular needs of a given tissue or developmental stage.In biochemistry, isozymes (or isoenzymes) are isoforms (closely related variants) of enzymes. In many cases, they are coded for by homologous genes that have diverged over time. Although, strictly speaking, allozymes represent enzymes from different alleles of the same gene, and isozymes represent enzymes from different genes that process or catalyse the same reaction, the two words are usually used interchangeably. Isozymes were first described by R. L. Hunter and Clement Markert (1957) who defined them as different variants of the same enzyme having identical functions and present in the same individual. This definition encompasses (1) enzyme variants that are the product of different genes and thus represent different loci (described as isozymes) and (2) enzymes that are the product of different alleles of the same gene (described as allozymes). Isozymes are usually the result of gene duplication, but can also arise from polyploidisation or nucleic acid hybridization. Over evolutionary time, if the function of the new variant remains identical to the original, then it is likely that one or the other will be lost as mutations accumulate, resulting in a pseudogene. However, if the mutations do not immediately prevent the enzyme from functioning, but instead modify either its function, or its pattern of expression, then the two variants may both be favoured by natural selection and become specialised to different functions. For example, they may be expressed at different stages of development or in different tissues. Allozymes may result from point mutations or from insertion-deletion (indel) events that affect the coding sequence of the gene. As with any other new mutations, there are three things that may happen to a new allozyme: An example of an isozyme is glucokinase, a variant of hexokinase which is not inhibited by glucose 6-phosphate. Its different regulatory features and lower affinity for glucose (compared to other hexokinases), allow it to serve different functions in cells of specific organs, such as control of insulin release by the beta cells of the pancreas, or initiation of glycogen synthesis by liver cells. Both these processes must only occur when glucose is abundant. The enzyme lactate dehydrogenase is a tetramer made of two different sub-units, the H-form and the M-form. These combine in different combinations depending on the tissue: Isozymes (and allozymes) are variants of the same enzyme. Unless they are identical in their biochemical properties, for example their substrates and enzyme kinetics, they may be distinguished by a biochemical assay. However, such differences are usually subtle, particularly between allozymes which are often neutral variants. This subtlety is to be expected, because two enzymes that differ significantly in their function are unlikely to have been identified as isozymes. While isozymes may be almost identical in function, they may differ in other ways. In particular, amino acid substitutions that change the electric charge of the enzyme are simple to identify by gel electrophoresis, and this forms the basis for the use of isozymes as molecular markers. To identify isozymes, a crude protein extract is made by grinding animal or plant tissue with an extraction buffer, and the components of extract are separated according to their charge by gel electrophoresis. Historically, this has usually been done using gels made from potato starch, but acrylamide gels provide better resolution. All the proteins from the tissue are present in the gel, so that individual enzymes must be identified using an assay that links their function to a staining reaction. For example, detection can be based on the localised precipitation of soluble indicator dyes such as tetrazolium salts which become insoluble when they are reduced by cofactors such as NAD or NADP, which generated in zones of enzyme activity. This assay method requires that the enzymes are still functional after separation (native gel electrophoresis), and provides the greatest challenge to using isozymes as a laboratory technique.