Function of tyrosine residues in rabbit muscle aldolase

doi:10.1016/0003-9861(67)90139-7

Archives of Biochemistry and Biophysics

Volume 122, Issue 1, October 1967, Pages 196-203

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Abstract

Acetylation of rabbit muscle aldolase with acetyl imidazole reduces the rate of cleavage of fructose diphosphate by 90%, but the rate of cleavage of fructose 1-phosphate is unaltered. The changes in catalytic properties are associated with acetylation of 25 of the 40 tyrosine residues in the protein. The COOH-terminal residues are not modified, although these changes resemble those observed when the enzyme is treated with carboxypeptidase. The enzyme-catalyzed incorporation of tritium from tritiated water into dihydroxyacetone phosphate is also impaired. In the presence of an acceptor aldehyde, such as acetaldehyde, the rate of cleavage of fructose diphosphate by the modified enzyme is increased. However, erythrose 4-phosphate, a natural substrate for aldolase, is not an acceptor. These results suggest that modification of tyrosine residues affects primarily the site binding the 6-phosphate of fructose diphosphate, and secondarily the ability of the Schiff base intermediate to dissociate from the enzyme.

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Fructose-1,6-diphosphate aldolase from Lactobacillus casei though a bacterial enzyme, shows functional similarities with rabbit muscle aldolase with respect to the involvement of amino acid residues for activity. Thus, in addition to lysine, which forms a Schiff base intermediate with the substrate, treatment of the enzyme with carboxypeptidase-A and tetranitromethane (TNM) suggested the presence of C-terminal tyrosine. Inactivation of the enzyme after exposure to light in the presence of rose Bengal indicated the presence of functional histidine residues. Finally, effects of thiol group reagents suggested involvement of these groups for maintaining the conformation essential for activity of the enzyme.
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Aldolase has been shown to be highly ubiquitous, occurring in all plant and animal tissues examined and in most microorganisms. It is absent in some heterofermentative bacteria such as Leuconostoc mesenteroides, and in some aerobic organisms such as Acetobacter suboxydans. On the basis of the difference in metal requirement, and other properties, aldolases have been classified into two categories—Class I and Class II. Class I aldolases, present in animals and higher plants, do not appear to require a metal ion cofactor and form a Schiff base intermediate with their substrate. These are inactivated by reduction with NaBH₄ in the presence of the substrate. The Class II aldolases, found in bacteria and molds, are not inactivated by reduction with NaBH₄ in the presence or absence of substrate. Certain blue-green algae appear to contain both types of aldolase. The molecular weight of Class II aldolases, where determined, is in the neighborhood of 70,000, approximately half that of mammalian aldolases, and they are composed of two, rather than four, subunits. The activity toward fructose-1, 6-bisphosphate (FDP) is not significantly affected by treatment with carboxypeptidase. Multiple forms of Class II aldolase have also been found in yeast and bacteria. Crystalline yeast aldolase can be resolved to yield three active components by isoelectric fractionation.
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^☆: This work was supported by grants from the National Institutes of Health (GM 11301) and the National Science Foundation (GB 1465). This is Communication No. 85 from the Joan and Lester Avnet Institute of Molecular Biology.

²: Postdoctoral fellow of the American Cancer Society (PF-308).

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Function of tyrosine residues in rabbit muscle aldolase☆

Abstract

Arch. Biochem. Biophys

J. Biol. Chem

Arch. Biochem. Biophys

J. Biol. Chem

J. Biol. Chem

J. Biol. Chem

J. Biol. Chem

Biochem. Biophys. Res. Commun

J. Biol. Chem

J. Biol. Chem

J. Biol. Chem

Arch. Biochem. Biophys