ISOTOPIC STUDIES OF ENZYME STRUCTURE AND FUNCTION
The understanding of enzyme action is not only fundamental to an elucidation of biological problems but also could lead to new developments in the important field of catalysis. The study of enzymes, however is complicated by the fragile nature of these compounds and the fact that only small amounts of material are usually available. Isotopic tehcniques have been found to be particularly useful in this field of investigation and the use of isotopes in two illustrative cases is described. The first example is the use of P/sup 32/to mark and then map out thc activc sitc of thc enzyme phosphoglucomutase. Enzymes are high moleculear weight compounds containing only a small area which is in direct contact with the substrate, To mark this site and onlu this site, radioactive substrates, glucose-1-P/sup 32/O/sub 4/ and glucose-6P/sup 32/O/sub 4/ , were prepared and incubated with enzyme until P/sup 32/-labelled enzyme was obtained. Tests using protein precipitants and inactive substrates demonstrated that this P/sup 32/ was covalently bound at the active site and that the labelled enxymen was completely active. Furthermore, it was shown that the labelled enzyme could be degraded to small peptides without migration or loss of the isotopic label. In this way it was found that 6 of ths amino acids at the active site are aspartic acid, serine, glycine, glutamic acid, alanine and valine. Moreover, nzymes suggests that a separation of amino acids into those involved in catalytic function and those involved in specifically function occurs in the protein structure. The second example involves the use a the C/sup 14/ -labelled methanol to elucidate the the role of wter in the proteins responsible for muscular contraction. Both myosin and a combination of actin and myosin, actomyosin catalyze the hydrolysis of adenosine triphosphate (ATP), but only the latter undergoes contraction By theoretical calculations it could be show that the existence and specificity of a water site on these proteins could be determined if methal phosphate in the range of 10/sup 12/ moles could be detected. This amount was well below the range of conventional analytical procedures but could be measured with the use of radioactive methanol. When the appropriate experiments were performed, it was found that both myosin and actomyosin have water sites and that these sites have similar specificities. The results indicate that the myosin part of actomyosin contains the locus at which water reacts during muscular contraction (auth)
- Research Organization:
- Brookhaven National Lab., Upton, N.Y.
- NSA Number:
- NSA-12-014572
- OSTI ID:
- 4318376
- Report Number(s):
- A/CONF.15/P/851
- Resource Relation:
- Other Information: Prepared for the Second U.N. International Conference on the Peaceful Uses of Atomic Energy, 1958. Orig. Receipt Date: 31-DEC-58
- Country of Publication:
- Country unknown/Code not available
- Language:
- English
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Related Subjects
ADENINE- ALCOHOLS- ATP- CHEMICAL REACTIONS- ENZYMES- HYDROLYSIS- LABELLED COMPOUNDS- MATHEMATICS- MEASURED VALUES- METHANOL- METHYL RADICALS- MUSCLES- MYOSIN- NUCLEIC ACIDS- ORGANIC COMPOUNDS- ORGANIC PHOSPHORUS COMPOUNDS- PHOSPHATES- PHYSIOLOGY- PROTEINS- QUANTITY RATIO- WATER
ALANINE- ALCOHOLS- AMINO ACIDS- ASPARTIC ACID- CARBON 14- CATALYSIS- ENZYMES- GLUTAMIC ACID- GLYCINE- - LABELLED COMPOUNDS- METHANOL- MUSCLES- PHYSIOLOGY- PROTEINS- SERINE- VALINE- WATER
CATALYSIS- CHEMICAL REACTIONS- CONFERENCE- DISTRIBUTION- ENZYMES- GLUCOSE- LABELLED COMPOUNDS- MOLECULES- PEPTIDES- PHOSPHATES- PHOSPHORU