YesT: a new rhamnogalacturonan acetyl esterase from Bacillus subtilis.
YesT, a putative protein from Bacillus subtilis ATCC 6633 that has been provisionally classified as a rhamnogalacturonan acetyl esterase (RGAE) in CE-12 family, was cloned, expressed in Escherichiacoli Rosetta (DE3), and purified. The enzyme is monomeric with a molecular mass of 37 kDa and presents thermophilic properties similar to RGAE from Aspergillus aculeatus, although YesT is more alkaliphilic. The study of inhibitors confirmed the importance of the His and the nucleophilic Ser for the esterase activity, apart from the Asp from the catalytic triad. This enzyme also presents broad substrate specificity, and is active toward 7-aminocephalosporanic acid, cephalosporin C, p-nitrophenyl acetate, beta-naphthyl acetate, glucose pentaacetate, and acetylated xylan. Moreover, YesT achieves a synergistic effect together with xylanase A toward acetylated xylan. As a member of the SGNH family, it does not adopt the common alpha/beta hydrolase fold. The primary sequence analysis and multiple sequence alignment revealed the lack of a two beta-stranded antiparallel sheet, which results in a clear change in the structure together with the disappearance of one of the three 3(10)-helices presented in RGAE structure. The similarities found in this article among the topological diagrams of RGAE, YesT, and Esterase A from Streptomyces scabies, Platelet-Activating Factor AcetylHydrolase, isoform Ib, alpha subunit [PAF-AH(Ib)alpha(1)], PAF-AH(Ib)alpha(2), the esterase domain from hemagglutinin esterase fusion glycoprotein (HEF1) from Influenza C virus, the thioesterase I (TAP) from E. coli, the hypothetical protein a1r1529 from Nostoc sp., and the hypothetical YxiM precursor that all belong to the SGNH family could indicate a possible divergence of such proteins from a common ancestor.
Martínez-Martínez
Irene
I
Faculty of Biology, Department of Biochemistry and Molecular Biology-A, University of Murcia, Campus Espinardo, E-30100 Murcia, Spain.
Navarro-Fernández
José
J
Daniel Lozada-Ramírez
José
J
García-Carmona
Francisco
F
Sánchez-Ferrer
Alvaro
A
D000115:Acetylesterase / Q000737:chemistry* / Q000145:classification; D001412:Bacillus subtilis / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000145:classification*; D011487:Protein Conformation
D001426:Bacterial Proteins; C096859:rhamnogalacturonan acetylesterase; D000115:Acetylesterase