Isozymes (also known as isoenzymes) are
enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters (i.e. different "K"M 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 (for example lactate dehydrogenase(LDH)).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 catalysethe same reaction, the two words are usually used interchangeably.
Isozymes were first described by
R. L. Hunterand 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 mutationsaccumulate, 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 gene expression, then the two variants may both be favoured by natural selectionand 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 DNAcoding sequence of the gene. As with any other new mutation, there are three things that may happen to a new allozyme:
# It is most likely that the new allele will be non-functional — in which case it will probably result in low fitness and be removed from the population by
# Alternatively, if the
amino acidresidue that is changed is in a relatively unimportant part of the enzyme, for example a long way from the active sitethen the mutation may be selectively neutral and subject to genetic drift.
# In rare cases the mutation may result in an enzyme that is more efficient, or one that can catalyse a slightly different
chemical reaction, in which case the mutation may cause an increase in fitness, and be favoured by natural selection.
An example of an isozyme
An example of an isozyme is
glucokinase, a variant of hexokinasewhich is not inhibited by glucose 6-phosphate. Its different regulatory features and lower affinity for glucose (compared to other hexokinases), allows it to serve different functions in cells of specific organs, such as control of insulinrelease by the beta cells of the pancreas, or initiation of glycogensynthesis by livercells. Both of these processes must only occur when glucose is abundant, or problems occur.
Isozymes (and allozymes) are variants of the same enzyme. Unless they are identical in terms of 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".
Whilst isozymes may be almost identical in function, they may differ in other ways. In particular,
amino acidsubstitutions that change the electric chargeof the enzyme (such as replacing aspartic acidwith glutamic acid) 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 potatostarch, however, acrylamidegels provide better resolution, and cellulose acetategels are now ( as of 2005) the norm.
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.
Isozymes and allozymes as molecular markers
Population geneticsis essentially a study of the causes and effects of genetic variation within and between populations, and in the past isozymes have been amongst the most widely used Molecular markers for this purpose. Although they have now been largely superseded by more informative DNA-based approaches (such as direct DNA sequencing, single nucleotide polymorphisms and microsatellites), they are still amongst the quickest and cheapest marker systems to develop, and remain ( as of 2005) an excellent choice for projects that only need to identify low levels of genetic variation, e.g. quantifying mating systems.
Other major examples
cytochrome P450isozymes play important roles in metabolismand steroidogenesis.
* Hunter, R. L. and C.L. Merkert. (1957) Histochemical demonstration of enzymes separated by zone electrophoresis in starch gels. "Science" 125: 1294-1295
* Wendel, JF, and NF Weeden. 1990. "Visualisation and interpretation of plant isozymes." Pp. 5-45 in D. E. Soltis and P. S. Soltis, eds. "Isozymes in plant biology." Chapman and Hall, London.
* Weeden, NF, and JF Wendel. 1990. "Genetics of plant isozymes". Pp. 46-72 in D. E. Soltis and P. S. Soltis, eds. "Isozymes in plant biology." Chapman and Hall, London
* Crawford, DJ. 1989. "Enzyme electrophoresis and plant systematics". Pp. 146-164 in D. E. Soltis and P. S. Soltis, eds. "Isozymes in plant biology." Dioscorides, Portland, Oregon.
*Hamrick, JL, and MJW Godt. 1990. "Allozyme diversity in plant species". Pp. 43-63 in A. H. D. Brown, M. T. Clegg, A. L. Kahler and B. S. Weir, eds. "Plant Population Genetics, Breeding, and Genetic Resources." Sinauer, Sunderland
* [http://www.genetics.cf.adfg.state.ak.us/techfac/electro2.php Allozyme Electrophoresis Techniques] - a complete guide to starch gel eletrophoresis
* [http://www.chem.ttu.ee/teadus/teema_en?t=T499 Development of new isozyme specific therapeutics] - Fatty Acid Dioxygenases and Eicosanoid Hormones (Estonia)
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