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Crystal structure of CTP‐ligated T state aspartate transcarbamoylase at 2.5 Å resolution: Implications for ATCase mutants and the mechanism of negative cooperativity
Author(s) -
Kosman Richard P.,
Gouaux J. Eric,
Lipscomb William N.
Publication year - 1993
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.340150206
Subject(s) - aspartate carbamoyltransferase , chemistry , allosteric regulation , stereochemistry , hydrogen bond , cooperativity , crystallography , side chain , active site , enzyme , molecule , biochemistry , organic chemistry , polymer
The X‐ray crystal structure of CTP‐ligated T state aspartate transcarbamoylase has been refined to an R factor of 0.182 at 2.5 Å resolution using the computer program X‐PLOR. The structure contains 81 sites for solvent and has rms deviations from ideality in bond lengths and bond angles of 0.018 Å and 3.722°, respectively. The cytosine base of CTP interacts with the main chain carbonyl oxygens of r Tyr‐89 and r Ile‐12, the main chain NH of r Ile‐12, and the amino group of r Lys‐60. The ribose hydroxyls form polar contacts with the amino group of r Lys‐60, a carboxylate oxygen of r Asp‐19, and the main chain carbonyl oxygen of r Val‐9. The phosphate oxygens of CTP interact with the amino group of r Lys‐94, the hydroxyl of r Thr‐82, and an imidazole nitrogen of r His‐20. Recent mutagenesis experiments evaluated in parallel with the structure reported here indicate that alterations in the hydrogen bonding environment of the side chain of r Asn‐111 may be responsible for the homotropic behavior of the pAR5 mutant of ATCase. The location of the first seven residues of the regulatory chain has been identified for the first time in a refined ATCase crystal structure, and the proximity of this portion of the regulatory chain to the allosteric site suggests a potential role for these residues in nucleotide binding to the enzyme. Finally, a series of amino acid side chain rearrangements leading from the R 1 CTP allosteric to the R 6 CTP allosteric site has been identified which may constitute the molecular mechanism of distinct CTP binding sites on ATCase. © 1993 Wiley‐Liss, Inc.