1.Structural insight into substrate specificity of human intestinal maltase-glucoamylase.
Limei REN ; Xiaohong QIN ; Xiaofang CAO ; Lele WANG ; Fang BAI ; Gang BAI ; Yuequan SHEN
Protein & Cell 2011;2(10):827-836
Human maltase-glucoamylase (MGAM) hydrolyzes linear alpha-1,4-linked oligosaccharide substrates, playing a crucial role in the production of glucose in the human lumen and acting as an efficient drug target for type 2 diabetes and obesity. The amino- and carboxyl-terminal portions of MGAM (MGAM-N and MGAM-C) carry out the same catalytic reaction but have different substrate specificities. In this study, we report crystal structures of MGAM-C alone at a resolution of 3.1 Å, and in complex with its inhibitor acarbose at a resolution of 2.9 Å. Structural studies, combined with biochemical analysis, revealed that a segment of 21 amino acids in the active site of MGAM-C forms additional sugar subsites (+ 2 and + 3 subsites), accounting for the preference for longer substrates of MAGM-C compared with that of MGAM-N. Moreover, we discovered that a single mutation of Trp1251 to tyrosine in MGAM-C imparts a novel catalytic ability to digest branched alpha-1,6-linked oligosaccharides. These results provide important information for understanding the substrate specificity of alpha-glucosidases during the process of terminal starch digestion, and for designing more efficient drugs to control type 2 diabetes or obesity.
Acarbose
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chemistry
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Amino Acid Sequence
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Catalytic Domain
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Crystallography, X-Ray
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Glycoside Hydrolase Inhibitors
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Humans
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Hydrogen Bonding
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Intestines
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enzymology
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Kinetics
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Maltose
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chemistry
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Molecular Sequence Data
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Mutagenesis, Site-Directed
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Mutation, Missense
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Oligosaccharides
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chemistry
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Pichia
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Protein Binding
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Recombinant Proteins
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antagonists & inhibitors
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chemistry
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genetics
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Substrate Specificity
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Surface Properties
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alpha-Glucosidases
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chemistry
;
genetics
2.The structure of WbnH in a near active state.
Fengzhi LI ; Siwei LI ; Xiaofen LIU ; Xue YANG ; Peng WANG ; Yuequan SHEN
Protein & Cell 2015;6(8):615-618
Binding Sites
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Catalytic Domain
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Crystallography, X-Ray
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Escherichia coli
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genetics
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metabolism
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Escherichia coli Proteins
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chemistry
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genetics
;
metabolism
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Models, Molecular
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N-Acetylgalactosaminyltransferases
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chemistry
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genetics
;
metabolism
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Protein Structure, Secondary
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Protein Structure, Tertiary
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Substrate Specificity
3.Crystal structures of catalytic core domain of BIV integrase: implications for the interaction between integrase and target DNA.
Xue YAO ; Shasha FANG ; Wentao QIAO ; Yunqi GENG ; Yuequan SHEN
Protein & Cell 2010;1(4):363-370
Integrase plays a critical role in the recombination of viral DNA into the host genome. Therefore, over the past decade, it has been a hot target of drug design in the fight against type 1 human immunodeficiency virus (HIV-1). Bovine immunodeficiency virus (BIV) integrase has the same function as HIV-1 integrase. We have determined crystal structures of the BIV integrase catalytic core domain (CCD) in two different crystal forms at a resolution of 2.45 Å and 2.2 Å, respectively. In crystal form I, BIV integrase CCD forms a back-to-back dimer, in which the two active sites are on opposite sides. This has also been seen in many of the CCD structures of HIV-1 integrase that were determined previously. However, in crystal form II, BIV integrase CCD forms a novel face-to-face dimer in which the two active sites are close to each other. Strikingly, the distance separating the two active sites is approximately 20 Å, a distance that perfectly matches a 5-base pair interval. Based on these data, we propose a model for the interaction of integrase with its target DNA, which is also supported by many published biochemical data. Our results provide important clues for designing new inhibitors against HIV-1.
Animals
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Catalytic Domain
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genetics
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Cattle
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DNA
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genetics
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DNA, Viral
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HIV-1
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genetics
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metabolism
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Humans
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Immunodeficiency Virus, Bovine
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enzymology
;
genetics
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Integrases
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chemistry
;
genetics
;
metabolism