Document Type

Article

Publication Date

2003

Journal or Book Title

Protein Engineering Design and Selection

Volume

16

Issue

7

First Page

521

Last Page

529

DOI

10.1093/protein/gzg066

Abstract

Aspergillus niger glucoamylase (GA) consists mainly of two forms, GAI [from the N‐terminus, catalytic domain + linker + starch‐binding domain (SBD)] and GAII (catalytic domain + linker). These domains were shuffled to make RGAI (SBD + linker + catalytic domain), RGAIΔL (SBD + catalytic domain) and RGAII (linker + catalytic domain), with domains defined by function rather than by tertiary structure. In addition, Paenibacillus macerans cyclomaltodextrin glucanotransferase SBD replaced the closely related A.niger GA SBD to give GAE. Soluble starch hydrolysis rates decreased as RGAII ≈ GAII ≈ GAI > RGAIΔL ≈ RGAI ≈ GAE. Insoluble starch hydrolysis rates were GAI > RGAIΔL > RGAI >> GAE ≈ RGAII > GAII, while insoluble starch‐binding capacities were GAI > RGAI > RGAIΔL > RGAII > GAII > GAE. These results indicate that: (i) moving the SBD to the N‐terminus or replacing the native SBD somewhat affects soluble starch hydrolysis; (ii) SBD location significantly affects insoluble starch binding and hydrolysis; (iii) insoluble starch hydrolysis is imperfectly correlated with its binding by the SBD; and (iv) placing the P.maceranscyclomaltodextrin glucanotransferase SBD at the end of a linker, instead of closely associated with the rest of the enzyme, severely reduces its ability to bind and hydrolyze insoluble starch.

Comments

This is a post-print of an article from Protein Engineering Design & Selection, 16, no. 7 (2003): 521–529, doi: 10.1093/protein/gzg066.

Copyright Owner

Oxford University Press

Language

en

File Format

application/pdf

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