Metabolic engineering of the morphology of Aspergillus oryzae by altering chitin synthesis.
Morphology and alpha-amylase production during submerged cultivation were examined in a wild-type strain (A1560) and in strains of Aspergillus oryzae in which chitin synthase B (chsB) and chitin synthesis myosin A (csmA) have been disrupted (ChsB/G and CM101). In a flowthrough cell, the growth of submerged hyphal elements was studied online, making it possible to examine the growth kinetics of the three strains. The average tip extension rates of the CM101 and ChsB/G strains were 25 and 88% lower, respectively, than that of the wild type. The branching intensity in the CM101 strain was 25% lower than that in the wild type, whereas that in the ChsB/G strain was 188% higher. During batch cultivation, inseparable clumps were formed in the wild-type strain, while no or fewer large inseparable clumps existed in the cultivations of the ChsB/G and CM101 strains. The alpha-amylase productivity was not significantly different in the three strains. A strain in which the transcription of chsB could be controlled by the nitrogen source-regulated promoter niiA (NiiA1) was examined during chemostat cultivation, and it was found that the branching intensity could be regulated by regulating the promoter, signifying an important role for chsB in branching. However, the pattern of branching responded very slowly to the change in transcription, and increased branching did not affect alpha-amylase productivity. alpha-Amylase residing in the cell wall was stained by immunofluorescence, and the relationship between tip number and enzyme secretion is discussed.
Müller
Christian
C
Center for Process Biotechnology, BioCentrum-DTU, Technical University of Denmark, 2800 Kgs. Lyngby.
McIntyre
Mhairi
M
Hansen
Kim
K
Nielsen
Jens
J
D001236:Aspergillus oryzae / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000648:ultrastructure*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002473:Cell Wall / Q000201:enzymology; D002686:Chitin / Q000378:metabolism*; D002687:Chitin Synthase / Q000235:genetics* / Q000378:metabolism*; D003470:Culture Media; D017353:Gene Deletion; D005818:Genetic Engineering / Q000379:methods*; D000516:alpha-Amylases / Q000378:metabolism
D001426:Bacterial Proteins; D003470:Culture Media; D002686:Chitin; D002687:Chitin Synthase; D000516:alpha-Amylases