3AAT

ACTIVITY AND STRUCTURE OF THE ACTIVE-SITE MUTANTS R386Y AND R386F OF ESCHERICHIA COLI ASPARTATE AMINOTRANSFERASE

Summary for 3AAT

• Entry DOI: 10.2210/pdb3aat/pdb
• Descriptor: ASPARTATE AMINOTRANSFERASE, SULFATE ION, PYRIDOXAL-5'-PHOSPHATE (3 entities in total)
• Functional Keywords: transferase(aminotransferase)
• Biological source: Escherichia coli
• Cellular location: Cytoplasm: P00509
• Total number of polymer chains: 1
• Total formula weight: 43952.40

Authors

Danishefsky, A.T.,Ringe, D.,Petsko, G.A. (deposition date: 1990-12-06, release date: 1992-01-15, Last modification date: 2024-06-05)

Primary citation

Danishefsky, A.T.,Onnufer, J.J.,Petsko, G.A.,Ringe, D.
Activity and structure of the active-site mutants R386Y and R386F of Escherichia coli aspartate aminotransferase.
Biochemistry, 30:1980-1985, 1991

PubMed Abstract: Arginine-386, the active-site residue of Escherichia coli aspartate aminotransferase (EC 2.6.1.1) that binds the substrate alpha-carboxylate, was replaced with tyrosine and phenylalanine by site-directed mutagenesis. This experiment was undertaken to elucidate the roles of particular enzyme-substrate interactions in triggering the substrate-induced conformational change in the enzyme. The activity and crystal structure of the resulting mutants were examined. The apparent second-order rate constants of both of these mutants are reduced by more than 5 orders of magnitude as compared to that of wild-type enzyme, though R386Y is slightly more active than R386F. The 2.5-A resolution structure of R386F in its native state was determined by using difference Fourier methods. The overall structure is very similar to that of the wild-type enzyme in the open conformation. The position of the Phe-386 side chain, however, appears to shift with respect to that of Arg-386 in the wild-type enzyme and to form new contacts with neighboring residues.

PubMed: 1993208
DOI: 10.1021/bi00221a035

Experimental method

X-RAY DIFFRACTION (2.8 Å)