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The use of natural and unnatural amino acid substrates to define the substrate specificity differences of escherichia coli aspartate and tyrosine aminotransferases
Author(s) -
Onuffer James J.,
Ton Binh T.,
Klement Ivan,
Kirsch Jack F.
Publication year - 1995
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560040909
Subject(s) - transamination , amino acid , stereochemistry , tyrosine , chemistry , side chain , enzyme , substrate (aquarium) , active site , aromatic amino acids , residue (chemistry) , carboxylate , alanine , biochemistry , biology , organic chemistry , ecology , polymer
The tyrosine (eTATase) and aspartate (eAATase) aminotransferases of Escherichia coli transaminate dicarboxylic amino acids with similar rate constants. However, eTATase exhibits ∼10 2 ‐10 4 ‐fold higher second‐order rate constants for the transamination of aromatic amino acids than does eAATase. A series of natural and unnatural amino acid substrates was used to quantitate specificity differences for these two highly related enzymes. A general trend toward lower transamination activity with increasing side‐chain length (extending from aspartate to glutamate to α ‐aminoadipate) is observed for both enzymes. This result suggests that dicarboxylate ligands associate with the two highly related enzymes in a similar manner. The high reactivity of the enzymes with L‐Asp and L‐Glu can be attributed to an ion pair interaction between the side‐chain carboxylate of the amino acid substrate and the guanidino group of the active site residue Arg 292 that is common to both enzymes. A strong linear correlation between side‐chain hydrophobicity and transamination rate constants obtains for n ‐alkyl side‐chain amino acid substrates with eTATase, but not for eAATase. The present kinetic data support a model in which eAATase contains one binding mode for all classes of substrate, whereas the active site of eTATase allows an additional mode that has increased affinity for hydrophobic amino acids.