1RPK

Crystal structure of barley alpha-amylase isozyme 1 (amy1) in complex with acarbose

Summary for 1RPK

• Entry DOI: 10.2210/pdb1rpk/pdb
• Related: 1AMY 1BG9 1HT6 1P6W 1RP8 1RP9
• Descriptor: Alpha-amylase type 1 isozyme, 4,6-dideoxy-4-{[(1S,5R,6S)-3-formyl-5,6-dihydroxy-4-oxocyclohex-2-en-1-yl]amino}-alpha-D-xylo-hex-5-enopyranose-(1-4)-alpha-D-glucopyranose, 4,6-dideoxy-4-{[(1S,5R,6S)-3-formyl-5,6-dihydroxy-4-oxocyclohex-2-en-1-yl]amino}-alpha-D-xylo-hex-5-enopyranose-(1-4)-alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose, ... (5 entities in total)
• Functional Keywords: alpha-amylase, barley, isozyme 1, beta-alpha-barrel, sugar tongs binding site, acarbose, hydrolase
• Biological source: Hordeum vulgare
• Cellular location: Secreted, extracellular space: P00693
• Total number of polymer chains: 1
• Total formula weight: 46354.68

Authors

Robert, X.,Haser, R.,Aghajari, N. (deposition date: 2003-12-03, release date: 2005-06-07, Last modification date: 2023-08-23)

Primary citation

Robert, X.,Haser, R.,Mori, H.,Svensson, B.,Aghajari, N.
Oligosaccharide Binding to Barley {alpha}-Amylase 1
J.Biol.Chem., 280:32968-32978, 2005

PubMed Abstract: Enzymatic subsite mapping earlier predicted 10 binding subsites in the active site substrate binding cleft of barley alpha-amylase isozymes. The three-dimensional structures of the oligosaccharide complexes with barley alpha-amylase isozyme 1 (AMY1) described here give for the first time a thorough insight into the substrate binding by describing residues defining 9 subsites, namely -7 through +2. These structures support that the pseudotetrasaccharide inhibitor acarbose is hydrolyzed by the active enzymes. Moreover, sugar binding was observed to the starch granule-binding site previously determined in barley alpha-amylase isozyme 2 (AMY2), and the sugar binding modes are compared between the two isozymes. The "sugar tongs" surface binding site discovered in the AMY1-thio-DP4 complex is confirmed in the present work. A site that putatively serves as an entrance for the substrate to the active site was proposed at the glycone part of the binding cleft, and the crystal structures of the catalytic nucleophile mutant (AMY1D180A) complexed with acarbose and maltoheptaose, respectively, suggest an additional role for the nucleophile in the stabilization of the Michaelis complex. Furthermore, probable roles are outlined for the surface binding sites. Our data support a model in which the two surface sites in AMY1 can interact with amylose chains in their naturally folded form. Because of the specificities of these two sites, they may locate/orient the enzyme in order to facilitate access to the active site for polysaccharide chains. Moreover, the sugar tongs surface site could also perform the unraveling of amylose chains, with the aid of Tyr-380 acting as "molecular tweezers."

PubMed: 16030022
DOI: 10.1074/jbc.M505515200

Experimental method

X-RAY DIFFRACTION (2 Å)