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Two novel, putatively cell wall-associated and glycosylphosphatidylinositol-anchored α-glucanotransferase enzymes of Aspergillus niger

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Author: Kaaij, R.M. van der · Yuan, X.L. · Franken, A. · Ram, A.F.J. · Punt, P.J. · Maarel, M.J.E.C. van der · Dijkhuizen, L.
Type:article
Date:2007
Institution: TNO Kwaliteit van Leven
Source:Eukaryotic Cell, 7, 6, 1178-1188
Identifier: 240062
doi: doi:10.1128/EC.00354-06
Keywords: Biology · Food technology · 4 alpha glucanotransferase · 4 alpha-glucanotransferase · fungal protein · glycogen debranching enzyme · glycosylphosphatidylinositol · hybrid protein · isoenzyme · maltooligosaccharides · oligosaccharide · unclassified drug · amino acid sequence · article · Aspergillus niger · cell wall · chemistry · classification · cytology · enzymology · genetics · metabolism · molecular genetics · nucleotide sequence · phylogeny · sequence alignment · sequence homology · Amino Acid Sequence · Aspergillus niger · Base Sequence · Cell Wall · Fungal Proteins · Glycogen Debranching Enzyme System · Glycosylphosphatidylinositols · Isoenzymes · Molecular Sequence Data · Oligosaccharides · Phylogeny · Recombinant Fusion Proteins · Sequence Alignment · Sequence Homology, Amino Acid · Aspergillus niger

Abstract

In the genome sequence of Aspergillus niger CBS 513.88, three genes were identified with high similarity to fungal α-amylases. The protein sequences derived from these genes were different in two ways from all described fungal α-amylases: they were predicted to be glycosylphosphatidylinositol anchored, and some highly conserved amino acids of enzymes in the α-amylase family were absent. We expressed two of these enzymes in a suitable A. niger strain and characterized the purified proteins. Both enzymes showed transglycosylation activity on donor substrates with α-(1,4)- glycosidic bonds and at least five anhydroglucose units. The enzymes, designated AgtA and AgtB, produced new α-(1,4)-glycosidic bonds and therefore belong to the group of the 4-α-glucanotransferases (EC 2.4.1.25). Their reaction products reached a degree of polymerization of at least 30. Maltose and larger maltooligosaccharides were the most efficient acceptor substrates, although AgtA also used small nigerooligosaccharides containing α-(1,3)-glycosidic bonds as acceptor substrate. An agtA knockout of A. niger showed an increased susceptibility towards the cell wall-disrupting compound calcofluor white, indicating a cell wall integrity defect in this strain. Homologues of AgtA and AgtB are present in other fungal species with α-glucans in their cell walls, but not in yeast species lacking cell wall α-glucan. Possible roles for these enzymes in the synthesis and/or maintenance of the fungal cell wall are discussed. Copyright © 2007, American Society for Microbiology. All Rights Reserved.