THE STRUCTURE OF ISOZYME SYSTEMS AND THEIR 

 ROLE IN DEVELOPMENT 



Edward J. Massaro 



Department of Biology, Yale University, New Haven, Connecticut 



In recent years, the study of isozymes has 

 expanded to such a degree that an attempt to 

 cover the field at a conference of this nature 

 would probably be only of minimal value. There- 

 fore, what I intend to do today is to use the 

 lactate dehydrogenase system of isozymes as a 

 model and present to you a fairly detailed view 

 of some of the work that has been and is being 

 pursued in this area. 



The individuality of cells, that is their 

 phenotype, is expressed in large measure by 

 the activitiesof their constituent enzymes. These 

 enzymes are the products of a complex series 

 of metabolic events that are under genetic con- 

 trol. In the broadest terms, enzyme biosynthesis 

 involves DNA transcription into RNA and RNA 

 translation into the linear amino acid sequence 

 or primary structure of a polypeptide chain. 

 Each polypeptide chain then assumes the charac- 

 teristic three dimensional conformation of its 

 secondary and tertiary structure. In numerous 

 instances these intricately folded polypeptide 

 chains are enzymatically inactive until they be- 

 come aggregated into more complex units (the 

 quarternary structure of the enzyme). 



The essence of the relationship between 

 DNA and enzyme structure has been summed-up 

 in the so-called one gene-one enzyme hypothesis. 

 A logical consequence of this hypothesis is that 

 the cells of a homozygous organism should 

 synthesize identical replicas of all of their con- 

 stituent protein molecules. But it has become 

 abundantly evident in recent years that numer- 

 ous proteins, including many enzymes, exist in 

 several physically distinctforms within the cells 

 of a single organism. The multiple molecular 

 forms of enxymes have been termed "iso- 

 zymes" (1). 



The isozymes of LDH exhibit both species- 

 and tissue-specific patterns (2). Furthermore, 

 during the course of embryonic development, 



these patterns undergo profound, albeit gradual, 

 changes (3). From such observations it seems 

 reasonable to propose that the remarkably 

 characteristic isozyme pattern of each tissue 

 reflects a physiological uniqueness of the indi- 

 vidual isozymes which is superimposed upon 

 their essential similarities. The existence of 

 isozymes then poses important questions con- 

 cerning their biosynthesis and their enzymatic 

 and physiological activities. 



LDH is ubiquitously distributed in isozy- 

 matic forms among vertebrates and also occurs 

 as such in numerous other organisms. It is an 

 oxido- reductase catalyzing the inter conversion 

 of lactate to pyruvate. This reaction is mediated 

 through the cofactor nicotinamide adenine di- 

 nucleotide (NAD). During periods of relative 

 anaerobiosis the enzyme functions to provide a 

 reservoir for the storage of hydrogen by form- 

 ing lactate which occupies a metabolic dead end. 

 This aids in maintaining the supply of NAD needed 

 at an earlier step in the glycolytic pathway. When 

 adequate supplies of NAD are again available, 

 lactate is oxidized to pyruvate. These reactions 

 are summarized in Fig. 1. It should be noted 

 that all the enzymes indicated on this chart, ex- 

 cept the three denoted by an asterisk, have been 

 shown to exist in multiple molecular forms. Re- 

 cent evidence indicates that triosephosphateiso- 

 merase also probably exists in isozymic forms. 



Although early investigations had un- 

 doubtedly demonstrated the existence of multiple 

 molecular forms of LDH, their biological sig- 

 nificance was not at first recognized (4, 5). In 

 1957 Vesell and Beam (6) described the existence 

 of three LDH isozymes in human serum and 

 changes in their proportions during various 

 disease states. About the same time, Wieland 

 and Pfleiderer (7) independently discovered the 

 existence of multiple molecular forms of LDH 

 and demonstrated tissue specific patterns of the 



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