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Identification of a trypsin-like serine protease from Trichoderma reesei QM9414

Paper ID Volume ID Publish Year Pages File Format Full-Text
18147 42713 2007 8 PDF Available
Title
Identification of a trypsin-like serine protease from Trichoderma reesei QM9414
Abstract

In the present work the genetically modified Trichoderma reesei strain QM9414 was used to produce full-length Cel7B (endoglucanase I, EGI) under the control of the constitutive Aspergillus nidulans gpdA promoter in the presence of glucose. However, the full-length Cel7B enzyme was found to be truncated to lower molecular weight components in the culture broth. Truncation of recombinant proteins produced in fungi may be due to protease activity. In order to identify major sectreted proteases, protease activity was assessed in culture filtrate of the T. reesei QM9414 recombinant. Zymogram analysis revealed the presence of proteolytic activity corresponding to one protein, which was subsequently purified by a combination of ion exchange and size exclusion chromatography. The protein has a molecular mass of 25 kDa, and an isoelectric point of 7.3. By matching tryptic peptide fragments analyzed by tandem mass spectrometry to fungal proteins available in databases as well as to expressed sequence tag (EST) sequences, and comparing the coded amino acids to full-length amino acid sequences, the purified protein was found to be homologous to several trypsin-like fungal serine proteases, with the highest homology to the protease P27 from Trichoderma harzianum. The purified protein was further characterized using benzoyl-arginyl-p-nitroanilide (BApNA) as substrate. It was found to have maximum activity at pH 8 and 50 °C, with a km-value of 0.3 mM.

Keywords
Trichoderma reesei; Cel7B (endoglucanase I); Serine protease; Trypsin
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Identification of a trypsin-like serine protease from Trichoderma reesei QM9414
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 40, Issue 5, 3 April 2007, Pages 1087–1094
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us