The noncatalytic triad of α-amylases: A novel structural motif involved in conformational stability

• Published in: Proteins, structure, function, and bioinformatics Vol. 70; no. 2; pp. 320 - 328
• Main Authors: Marx, Jean-Claude, Poncin, Johan, Simorre, Jean-Pierre, Ramteke, Pramod W., Feller, Georges
• Format: Journal Article Web Resource
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01-02-2008; Wiley-Liss Inc
• Subjects: alpha-Amylases - chemistry; alpha-Amylases - genetics; alpha-Amylases - metabolism; Biochemistry, biophysics & molecular biology; Biochimie, biophysique & biologie moléculaire; Catalysis; catalytic triad; Enzyme Stability; Life sciences; low barrier hydrogen bond; Mutagenesis, Site-Directed; Nuclear Magnetic Resonance, Biomolecular; protease; Protein Conformation; protein stability; Pseudoalteromonas - enzymology; Sciences du vivant; α-amylase
• ISSN: 0887-3585; 1097-0134; 1097-0134
• DOI: 10.1002/prot.21594

Chloride‐activated α‐amylases contain a noncatalytic triad, independent of the glycosidic active site, perfectly mimicking the catalytic triad of serine‐proteases and of other active serine hydrolytic enzymes. Mutagenesis of Glu, His, and Ser residues in various α‐amylases shows that this pattern is a structural determinant of the enzyme conformation that cannot be altered without losing the intrinsic stability of the protein. 1H‐15N NMR spectra of a bacterial α‐amylase reveal proton signals that are identical with the NMR signature of catalytic triads and especially a deshielded proton involving a protonated histidine and displaying properties similar to that of a low barrier hydrogen bond. It is proposed that the H‐bond between His and Glu of the noncatalytic triad is an unusually strong interaction, responsible for the observed NMR signal and for the weak stability of the triad mutants. Furthermore, a stringent template‐based search of the Protein Data Bank demonstrated that this motif is not restricted to α‐amylases, but is also found in 80 structures from 33 different proteins, amongst which SH2 domain‐containing proteins are the best representatives. Proteins 2008. © 2007 Wiley‐Liss, Inc.