2D0H

Crystal Structure of Thermoactinomyces vulgaris R-47 Alpha-Amylase 1 (TVAI) Mutant D356N/E396Q complexed with P2, a pullulan model oligosaccharide

Summary for 2D0H

• Entry DOI: 10.2210/pdb2d0h/pdb
• Related: 1JI1 2D0F 2D0G
• Descriptor: alpha-amylase I, alpha-D-glucopyranose-(1-6)-alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose-(1-6)-alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose, CALCIUM ION, ... (5 entities in total)
• Functional Keywords: alpha-amylase, hydrolase
• Biological source: Thermoactinomyces vulgaris
• Cellular location: Secreted: Q60053
• Total number of polymer chains: 1
• Total formula weight: 72337.60

Authors

Abe, A.,Yoshida, H.,Tonozuka, T.,Sakano, Y.,Kamitori, S. (deposition date: 2005-08-02, release date: 2006-07-11, Last modification date: 2024-05-29)

Primary citation

Abe, A.,Yoshida, H.,Tonozuka, T.,Sakano, Y.,Kamitori, S.
Complexes of Thermoactinomyces vulgaris R-47 alpha-amylase 1 and pullulan model oligossacharides provide new insight into the mechanism for recognizing substrates with alpha-(1,6) glycosidic linkages
Febs J., 272:6145-6153, 2005

PubMed Abstract: Thermoactinomyces vulgaris R-47 alpha-amylase 1 (TVAI) has unique hydrolyzing activities for pullulan with sequence repeats of alpha-(1,4), alpha-(1,4), and alpha-(1,6) glycosidic linkages, as well as for starch. TVAI mainly hydrolyzes alpha-(1,4) glycosidic linkages to produce a panose, but it also hydrolyzes alpha-(1,6) glycosidic linkages with a lesser efficiency. X-ray structures of three complexes comprising an inactive mutant TVAI (D356N or D356N/E396Q) and a pullulan model oligosaccharide (P2; [Glc-alpha-(1,6)-Glc-alpha-(1,4)-Glc-alpha-(1,4)]2 or P5; [Glc-alpha-(1,6)-Glc-alpha-(1,4)-Glc-alpha-(1,4)]5) were determined. The complex D356N/P2 is a mimic of the enzyme/product complex in the main catalytic reaction of TVAI, and a structural comparison with Aspergillus oryzaealpha-amylase showed that the (-) subsites of TVAI are responsible for recognizing both starch and pullulan. D356N/E396Q/P2 and D356N/E396Q/P5 provided models of the enzyme/substrate complex recognizing the alpha-(1,6) glycosidic linkage at the hydrolyzing site. They showed that only subsites -1 and -2 at the nonreducing end of TVAI are effective in the hydrolysis of alpha-(1,6) glycosidic linkages, leading to weak interactions between substrates and the enzyme. Domain N of TVAI is a starch-binding domain acting as an anchor in the catalytic reaction of the enzyme. In this study, additional substrates were also found to bind to domain N, suggesting that domain N also functions as a pullulan-binding domain.

PubMed: 16302977
DOI: 10.1111/j.1742-4658.2005.05013.x

Experimental method

X-RAY DIFFRACTION (2.1 Å)