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Titolo:
Design, X-ray crystallography, molecular modelling and thermal stability studies of mutant enzymes at site 172 of 3-isopropylmalate dehydrogenase from Thermus thermophilus
Autore:
Qu, CX; Akanuma, S; Tanaka, N; Moriyama, H; Ohsima, T;
Indirizzi:
Tokyo Inst Technol, Grad Sch Biosci & Biotechnol, Dept Life Sci, Yokohama,Kanagawa 2268501, Japan Tokyo Inst Technol Yokohama Kanagawa Japan 2268501anagawa 2268501, Japan Tokyo Univ Pharm & Life Sci, Dept Mol Biol, Tokyo 1920392, Japan Tokyo Univ Pharm & Life Sci Tokyo Japan 1920392 ol, Tokyo 1920392, Japan
Titolo Testata:
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
, volume: 57, anno: 2001,
parte:, 2
pagine: 225 - 232
SICI:
0907-4449(200102)57:<225:DXCMMA>2.0.ZU;2-8
Fonte:
ISI
Lingua:
ENG
Soggetto:
EXTREME THERMOPHILE; HYDROPHOBIC CORE; T4 LYSOZYME; BACTERIOPHAGE-T4 LYSOZYME; CONFORMATIONAL STABILITY; THERMOSTABLE MUTANTS; STRUCTURAL-ANALYSIS; PROTEIN STRUCTURES; CRYSTAL-STRUCTURES; BACILLUS-SUBTILIS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
38
Recensione:
Indirizzi per estratti:
Indirizzo: Tanaka, N Tokyo Inst Technol, Grad Sch Biosci & Biotechnol, Dept Life Sci,Nagatsuta4259, Yokohama, Kanagawa 2268501, Japan Tokyo Inst Technol Nagatsuta 4259 Yokohama Kanagawa Japan 2268501
Citazione:
C.X. Qu et al., "Design, X-ray crystallography, molecular modelling and thermal stability studies of mutant enzymes at site 172 of 3-isopropylmalate dehydrogenase from Thermus thermophilus", ACT CRYST D, 57, 2001, pp. 225-232

Abstract

The relationship between the structure and the thermostability of the 3-isopropylmalate dehydrogenase from Thermus thermophilus was studied by site-directed mutation of a single Ala residue located at the domain interface. The crystal structures of three mutant enzymes, replacing Ala172 with Gly, Val and Phe, were successfully determined at 2.3, 2.2 and 2.5 Angstrom resolution, respectively. Substitution of Ala172 by relatively 'short' residues (Gly, Val or Ile) enlarges or narrows the cavity in the vicinity of the C-beta atom of Ala172 and the thermostability of the enzyme shows a good correlation with the hydrophobicity of the substituted residues. Substitution ofAla172 by the 'longer' residues Leu or Phe causes a rearrangement of the domain structure, which leads to a higher thermostability of the enzymes than that expected from the hydrophobicity of the substituted residues. Mutation of Ala172 to negatively charged residues gave an unexpected result: the melting temperature of the Asp mutant enzyme was reduced by 2.7 K while that of the Glu mutant increased by 1.8 K. Molecular-modelling studies indicated that the glutamate side chain was sufficiently long that it did not act as a buried charge as did the aspartate, but instead protruded to the outside of the hydrophobic cavity and contributed to the stability of the enzymeby enhancing the packing of the local side chains and forming an extra salt bridge with the side chain of Lys175.

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Documento generato il 05/12/20 alle ore 12:34:37