Hen egg white lysozyme expressed in, and secreted from, Aspergillus niger is correctly processed and folded.
We transformed Aspergillus niger with the full length cDNA gene encoding hen egg-white lysozyme (HEWL) and its secretion signal sequence. Lysozyme levels up to 12 mg/l were secreted when expression was controlled by the A. awamori glucoamylase (GAM) promoter and 1 mg/l when controlled by the A. nidulans glyceraldehyde-3-phosphate dehydrogenase (GPD) promoter. N-terminal sequence analysis of the recombinant protein indicated that the signal peptide was correctly processed by the A. niger secretory apparatus. The specific catalytic activity of the recombinant protein was identical to that of authentic hen lysozyme. The recombinant HEWL was examined by 2D 1H-NMR spectroscopy and shown to have a spectrum identical to that of authentic HEWL indicating that the protein was correctly folded.
Archer
D B
DB
Agricultural and Food Research Council Institute of Food Research, Norwich Laboratory, U.K.
Jeenes
D J
DJ
MacKenzie
D A
DA
Brightwell
G
G
Lambert
N
N
Lowe
G
G
Radford
S E
SE
Dobson
C M
CM
D000595:Amino Acid Sequence; D000818:Animals; D001234:Aspergillus niger / Q000201:enzymology / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D005087:Glucan 1,4-alpha-Glucosidase / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009113:Muramidase / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D010063:Ovum / Q000201:enzymology; D010957:Plasmids; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism; D014162:Transfection / Q000235:genetics
D011993:Recombinant Fusion Proteins; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D009113:Muramidase; D005087:Glucan 1,4-alpha-Glucosidase