Phylogenetic and biochemical characterization of a novel cluster of intracellular fungal alpha-amylase enzymes.
Currently known fungal alpha-amylases are well-characterized extracellular enzymes that are classified into glycoside hydrolase subfamily GH13_1. This study describes the identification, and phylogenetic and biochemical analysis of novel intracellular fungal alpha-amylases. The phylogenetic analysis shows that they cluster in the recently identified subfamily GH13_5 and display very low similarity to fungal alpha-amylases of family GH13_1. Homologues of these intracellular enzymes are present in the genome sequences of all filamentous fungi studied, including ascomycetes and basidiomycetes. One of the enzymes belonging to this new group, Amy1p from Histoplasma capsulatum, has recently been functionally linked to the formation of cell wall alpha-glucan. To study the biochemical characteristics of this novel cluster of alpha-amylases, we overexpressed and purified a homologue from Aspergillus niger, AmyD, and studied its activity product profile with starch and related substrates. AmyD has a relatively low hydrolysing activity on starch (2.2 U mg(-1)), producing mainly maltotriose. A possible function of these enzymes in relation to cell wall alpha-glucan synthesis is discussed.
van der Kaaij
R M
RM
Centre for Carbohydrate Bioprocessing, TNO-University of Groningen, Haren, The Netherlands.|Microbial Physiology Research Group, Groningen Biomolecular Sciences and Biotechnology Institute (GBB), University of Groningen, The Netherlands.
Janeček
Š
Š
Institute of Molecular Biology, Slovak Academy of Sciences, Bratislava, Slovakia.
van der Maarel
M J E C
MJEC
Business Unit Food and Biotechnology Innovations, TNO Quality of Life, Groningen, The Netherlands.|Centre for Carbohydrate Bioprocessing, TNO-University of Groningen, Haren, The Netherlands.|Microbial Physiology Research Group, Groningen Biomolecular Sciences and Biotechnology Institute (GBB), University of Groningen, The Netherlands.
Dijkhuizen
L
L
Centre for Carbohydrate Bioprocessing, TNO-University of Groningen, Haren, The Netherlands.|Microbial Physiology Research Group, Groningen Biomolecular Sciences and Biotechnology Institute (GBB), University of Groningen, The Netherlands.
D000595:Amino Acid Sequence; D000818:Animals; D001234:Aspergillus niger / Q000201:enzymology / Q000235:genetics; D002473:Cell Wall / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005658:Fungi / Q000737:chemistry / Q000145:classification / Q000201:enzymology* / Q000235:genetics*; D005936:Glucans / Q000378:metabolism; D006658:Histoplasma / Q000201:enzymology / Q000235:genetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D016415:Sequence Alignment; D013213:Starch / Q000378:metabolism; D013379:Substrate Specificity; D000516:alpha-Amylases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D005656:Fungal Proteins; D005936:Glucans; D013213:Starch; D000516:alpha-Amylases