Properties of N‐terminus truncated and C‐terminus mutated muscle acylphosphatases

FEBS Letters

Raugei, Giovanni¹  Stefani, Massimo¹  Magherini, Francesca¹  Taddei, Niccolò¹  Bucciantini, Monica¹  Modesti, Alessandra¹  Ramponi, Giampietro¹  Vecchi, Manuela¹ 

[1] Department of Biochemical Sciences, University of Florence, V.le Morgagni 50, 50134 Florence, Italy

关键词: Acylphosphatase deletion mutant;    Acylphosphatase recombinant;    Acylphosphatase mutant;    Acylphosphatase 1H NMR spectrum;    Y98Q;    tyrosine 98 to glutamine mutant;    R97Q;    arginine 97 to glutamine mutant;    Δ6;    lacking residues 1–6 at the N-terminus;    IPTG;    isopropyl-thiogalactoside;    CED-;    cyanoethyldeoxy-;    NMR;    nuclear magnetic resonance;   

DOI  :  10.1016/0014-5793(95)00236-3

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Enzymatic activity and structure of N-terminus truncated and C-terminus substituted muscle acylphosphatase mutants were investigated by kinetic studies under different conditions and ¹H NMR spectroscopy, respectively. The N-terminus truncated mutant lacked the first six residues (Δ6), whereas arginine 97 and tyrosine 98 were replaced by glutamine giving two C-terminus substituted mutants (R97Q and Y98Q, respectively). All acylphosphatase forms were obtained by modifications of a synthetic gene coding for the human muscle enzyme which was expressed in E. coli. The Δ6 deletion mutant elicited a reduced specific activity and a native-like structure. The kinetic and structural properties of R97Q and Y98Q mutants indicate a possible role of Arg-97 in the stabilisation of the active site correct conformation, most likely via back-bone and side chain interactions with Arg-23, the residue involved in phosphate binding by the enzyme. This study also suggests a possible involvement of Tyr-98 in the stabilisation of the acylphosphatase overall structure.

【 授权许可】

Unknown