title: Crystal structure and thermostability of a putative α-glucosidase from Thermotoga neapolitana.
abstract: Glycoside hydrolase family 4 (GH4) represents an unusual group of glucosidases with a requirement for NAD(+), Mn(2+), and reducing conditions. We found a putative α-glucosidase belonging to GH4 in hyperthermophilic Gram-negative bacterium Thermotoga neapolitana. In this study, we recombinantly expressed the putative α-glycosidase from T. neapolitana, and determined the crystal structure of the protein at a resolution of 2.0Å in the presence of Mn(2+) but in the absence of NAD(+). The structure showed the dimeric assembly and the Mn(2+) coordination that other GH4 enzymes share. In comparison, we observed structural changes in T. neapolitana α-glucosidase by the binding of NAD(+), which also increased the thermostability. Numerous arginine-mediated salt-bridges were observed in the structure, and we confirmed that the salt bridges correlated with the thermostability of the proteins. Disruption of the salt bridge that linked N-terminal and C-terminal parts at the surface dramatically decreased the thermostability. A mutation that changed the internal salt bridge to a hydrogen bond also decreased the thermostability of the protein. This study will help us to understand the function of the putative glucosidase and the structural features that affect the thermostability of the protein.
authors.affiliation: Department of Manufacturing Pharmacy, College of Pharmacy and Research Institute for Drug Development, Pusan National University, Geumjeong-gu, Busan 609-735, Republic of Korea.
authors.forename: Bo-Young
authors.initials: BY
authors.lastname: Yun
authors.forename: So-Young
authors.initials: SY
authors.lastname: Jun
authors.forename: Nam Ah
authors.initials: NA
authors.lastname: Kim
authors.forename: Bo-Young
authors.initials: BY
authors.lastname: Yoon
authors.forename: Shunfu
authors.initials: S
authors.lastname: Piao
authors.forename: So-Hae
authors.initials: SH
authors.lastname: Park
authors.forename: Seong Hoon
authors.initials: SH
authors.lastname: Jeong
authors.forename: Heeseob
authors.initials: H
authors.lastname: Lee
authors.forename: Nam-Chul
authors.initials: NC
authors.lastname: Ha
mesh_terms: D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D004795:Enzyme Stability; D006358:Hot Temperature; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D046949:Thermotoga neapolitana / Q000201:enzymology*; D000520:alpha-Glucosidases / Q000737:chemistry* / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D000520:alpha-Glucosidases