Advanced Search

Journal Navigation

Journal Home

Subscriptions

Archive

Contact Us

Table of Contents

Sign In to gain access to subscriptions and/or personal tools.
Journal of Endotoxin Research
This Article
Right arrow Full Text (PDF)
Right arrow References
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Alert me to new issues of the journal
Right arrow Add to Saved Citations
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Request Reprints
Right arrow Add to My Marked Citations
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Right arrow Citing Articles via Scopus
Google Scholar
Right arrow Articles by Ding, J.L.
Right arrow Articles by Ho, B.
Right arrow Search for Related Content
PubMed
Right arrow Articles by Ding, J.L.
Right arrow Articles by Ho, B.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

Expression of full length and deletion homologues of Carcinoscorpius rotundicauda Factor C in Saccharomyces cerevisiae: immunoreactivity and endotoxin binding

J.L. Ding

Marine Biotechnology Laboratory and BioScience Centre, sbsdjl{at}leonis.nus.sg

C. Chai

Department of Microbiology, School of Biological Sciences National University of Singapore, Singapore

A.W.M. Pui

Marine Biotechnology Laboratory and BioScience Centre

B. Ho

Department of Microbiology, School of Biological Sciences National University of Singapore, Singapore

Deletion homologues of the cloned Factor C cDNAs from the horseshoe crab Carcinoscorpius rotundicauda were engineered to express in Saccharomyces cerevisiae under the regulation of a galactose-inducible promoter. Expression cassettes were constructed in the vectors: pEMBLyex4 and YEpsec1 to direct, respectively, the intracellular expression, and the secretion of the protein into the culture medium using a heterologous signal sequence. The effect of insert size on the efficiency of expression and the functionality of the resulting recombinant Factor C (rFC) were studied by creating expression constructs bearing various deletion and/or hybrid fragments of Factor C. Removal of the long 5' UTR from the Factor C cDNA improved expression of the rFC. 3' Deletions of up to 84%, or internal deletions of 65% of the Factor C cDNA resulted in either the lack of detectable amounts of Factor C or loss of immunoreactivity. Depending on the construct, full length or partial rFC-related proteins were correspondingly expressed intracellularly, regardless of the vector. The rFC partitioned with the insoluble cell fraction, was solubilised with either SDS or Triton X-100, and found to be immunoreactive. The rFCs were functionally active, being able to bind Gram-negative bacterial endotoxin, provided critical regions of the endotoxin-binding domain were preserved.

Journal of Endotoxin Research, Vol. 4, No. 1, 33-43 (1997)
DOI: 10.1177/096805199700400105
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?


This article has been cited by other articles:


Home page
 Antimicrob. Agents Chemother.Home page
Y. H. Yau, B. Ho, N. S. Tan, M. L. Ng, and J. L. Ding
High Therapeutic Index of Factor C Sushi Peptides: Potent Antimicrobials against Pseudomonas aeruginosa
Antimicrob. Agents Chemother., October 1, 2001; 45(10): 2820 - 2825.
[Abstract] [Full Text]