Degradation of Granular Starch by the Bacterium Microbacterium aurum Strain B8.A Involves a Modular α-Amylase Enzyme System with FNIII and CBM25 Domains

The bacterium Microbacterium aurum strain B8.A, originally isolated from a potato plant wastewater facility, is able to degrade different types of starch granules. Here we report the characterization of an unusually large, multidomain M. aurum B8.A α-amylase enzyme (MaAmyA). MaAmyA is a 1,417-amino-...

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Bibliographic Details
Published in:Applied and Environmental Microbiology 2015-10, Vol.81 (19), p.6610-6620
Main Authors: Valk, Vincent, Eeuwema, Wieger, Sarian, Fean D, van der Kaaij, Rachel M, Dijkhuizen, Lubbert
Format: Article
Language:English
Subjects:
Actinomycetales - chemistry
Actinomycetales - classification
Actinomycetales - enzymology
Actinomycetales - genetics
alpha-Amylases - chemistry
alpha-Amylases - genetics
alpha-Amylases - metabolism
Bacteria
Bacterial Proteins - chemistry
Bacterial Proteins - genetics
Bacterial Proteins - metabolism
Biocatalysis
Enzymology and Protein Engineering
Hydrolysis
Microbacterium
Molecular Sequence Data
Phylogeny
Protein Structure, Tertiary
Solanum tuberosum
Solanum tuberosum - metabolism
Starch - metabolism
Triticum aestivum
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Summary:The bacterium Microbacterium aurum strain B8.A, originally isolated from a potato plant wastewater facility, is able to degrade different types of starch granules. Here we report the characterization of an unusually large, multidomain M. aurum B8.A α-amylase enzyme (MaAmyA). MaAmyA is a 1,417-amino-acid (aa) protein with a predicted molecular mass of 148 kDa. Sequence analysis of MaAmyA showed that its catalytic core is a family GH13_32 α-amylase with the typical ABC domain structure, followed by a fibronectin (FNIII) domain, two carbohydrate binding modules (CBM25), and another three FNIII domains. Recombinant expression and purification yielded an enzyme with the ability to degrade wheat and potato starch granules by introducing pores. Characterization of various truncated mutants of MaAmyA revealed a direct relationship between the presence of CBM25 domains and the ability of MaAmyA to form pores in starch granules, while the FNIII domains most likely function as stable linkers. At the C terminus, MaAmyA carries a 300-aa domain which is uniquely associated with large multidomain amylases; its function remains to be elucidated. We concluded that M. aurum B8.A employs a multidomain enzyme system to initiate degradation of starch granules via pore formation.
ISSN:0099-2240
1098-5336
1098-6596
DOI:10.1128/AEM.01029-15