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Isolation and Characterization of a Proteinaceous α-amylase Inhibitor AAI-CC5 From Streptomyces Sp. CC5, and Its Gene Cloning and Expression

Isolation and Characterization of a Proteinaceous α-amylase Inhibitor AAI-CC5 From Streptomyces Sp. CC5, and Its Gene Cloning and Expression

Zhibin Sun, Weihao Lu, Pingping Liu, Hui Wang, Yan Huang, Yuguo Zhao, Yi Kong, Zhongli Cui

Antonie Van Leeuwenhoek. 2015 Feb;107(2):345-56.

PMID: 25411086

Abstract:

An α-amylase inhibitor producing Streptomyces sp. strain CC5 was isolated from soil. A proteinaceous α-amylase inhibitor AAI-CC5 was purified from strain CC5. AAI-CC5 specifically inhibited mammalian α-amylases. The molecular weight of the inhibitor was determined to be 8,212 Da by MALDI-TOF Mass Spectrum. The N-terminal 15 amino acid residues of the purified AAI-CC5 were DTGSPAPECVEYFQS, which is dissimilar to other reported proteinaceous α-amylase inhibitors. AAI-CC5 is a pH insensitive and heat-stable protein, and cannot be hydrolysed by trypsin. AAI-CC5 was cloned and expressed in Escherichia coli BL21 (DE3) with a hexa-histidine tag on the C terminal. AAI-CC5 shared 82 % identity with Parvulustat. The recombinant α-amylase inhibitor was purified to homogeneity by one-step affinity chromatography using Ni(2+)-NTA resin with molecular mass of 9,404 Da. Steady state kinetics studies of α-amylase and the inhibitor revealed an irreversible, non-competitive inhibition mechanism with IC50 and Ki value of 6.43 ×1 10(-11) and 4.45 × 10(-11) M respectively. These results suggest this novel α-amylase inhibitor possessed powerful inhibitory activity for α-amylase, and it may be a candidate in research of diabetes therapy and obesity treatment.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP9000855-A	α-Amylase, heat-stable		9000-85-5	Price

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