title: Structure of the thermophilic l-Arabinose isomerase from Geobacillus kaustophilus reveals metal-mediated intersubunit interactions for activity and thermostability.
abstract: Thermophilic l-arabinose isomerase (AI), which catalyzes the interconversion of l-arabinose and l-ribulose, can be used to produce d-tagatose, a sugar substitute, from d-galactose. Unlike mesophilic AIs, thermophilic AIs are highly dependent on divalent metal ions for their catalytic activity and thermostability at elevated temperatures. However, the molecular basis underlying the substrate preferences and metal requirements of multimeric AIs remains unclear. Here we report the first crystal structure of the apo and holo forms of thermophilic Geobacillus kaustophilus AI (GKAI) in hexamer form. The structures, including those of GKAI in complex with l-arabitol, and biochemical analyses revealed not only how the substrate-binding site of GKAI is formed through displacement of residues at the intersubunit interface when it is bound to Mn(2+), but also revealed the water-mediated H-bonding networks that contribute to the structural integrity of GKAI during catalysis. These observations suggest metal-mediated isomerization reactions brought about by intersubunit interactions at elevated temperatures are responsible for the distinct active site features that promote the substrate specificity and thermostability of thermophilic AIs.
authors.affiliation: Department of Cellular and Molecular Medicine, Chosun University School of Medicine, Gwangju, 61452, South Korea.
authors.forename: Jin Myung
authors.initials: JM
authors.lastname: Choi
authors.affiliation: School of Applied Biosciences, Kyungpook National University, Daegu, 41566, South Korea.
authors.forename: Yong-Jik
authors.initials: YJ
authors.lastname: Lee
authors.affiliation: Department of Cellular and Molecular Medicine, Chosun University School of Medicine, Gwangju, 61452, South Korea.
authors.forename: Thinh-Phat
authors.initials: TP
authors.lastname: Cao
authors.affiliation: School of Applied Biosciences, Kyungpook National University, Daegu, 41566, South Korea.
authors.forename: Sun-Mi
authors.initials: SM
authors.lastname: Shin
authors.affiliation: School of Applied Biosciences, Kyungpook National University, Daegu, 41566, South Korea.
authors.forename: Min-Kyu
authors.initials: MK
authors.lastname: Park
authors.affiliation: Major in Food Biotechnology, Silla University, Busan, 46958, South Korea.
authors.forename: Han-Seung
authors.initials: HS
authors.lastname: Lee
authors.affiliation: School of Life Sciences, Kyungpook National University, Daegu, 41566, South Korea.
authors.forename: Eric
authors.initials: E
authors.lastname: di Luccio
authors.affiliation: CJ CheilJedang, Life Ingredient & Material Research Institute, Suwon, 16495, South Korea.
authors.forename: Seong-Bo
authors.initials: SB
authors.lastname: Kim
authors.affiliation: Major in Food Biotechnology, Silla University, Busan, 46958, South Korea.
authors.forename: Sang-Jae
authors.initials: SJ
authors.lastname: Lee
authors.affiliation: Microbiomics & Immunity Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, South Korea.
authors.forename: Sang Jun
authors.initials: SJ
authors.lastname: Lee
authors.affiliation: Department of Cellular and Molecular Medicine, Chosun University School of Medicine, Gwangju, 61452, South Korea. Electronic address: sunglee@chosun.ac.kr.
authors.forename: Sung Haeng
authors.initials: SH
authors.lastname: Lee
authors.affiliation: School of Applied Biosciences, Kyungpook National University, Daegu, 41566, South Korea. Electronic address: leehicam@knu.ac.kr.
authors.forename: Dong-Woo
authors.initials: DW
authors.lastname: Lee
mesh_terms: D019747:Aldose-Ketose Isomerases / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray; D004795:Enzyme Stability; D056506:Geobacillus / Q000201:enzymology*; D006358:Hot Temperature; D006860:Hydrogen Bonding; D008345:Manganese / Q000737:chemistry*; D020836:Protein Structure, Quaternary
chemical_list: D001426:Bacterial Proteins; D008345:Manganese; D019747:Aldose-Ketose Isomerases; C019715:L-arabinose isomerase
keywords: Crystal structure; Intersubunit interaction; Substrate specificity; Thermostability; l-arabinose isomerase