Cloned factor C cDNA of the Singapore horseshoe crab, Carcinoscorpius rotundicauda and purification of factor C proenzyme
US5716834A · kind A · utility
Assignee
Inventors
Key dates
| Filing date | Aug 19, 1994 |
| Grant date | Feb 10, 1998 |
| Priority date | — |
| Expiry date | Aug 19, 2014 |
Classification
- Technology area (CPC G)Physics
- CPC primaryG01N33/579
- WIPO fieldMeasurement
- WIPO sectorInstruments
Abstract
Full-length and deletion subclones of cDNAs for Factor C of Carcinoscorpius rotundicauda are provided. These cDNAs have been cloned into .lambda.gt 22 and pGEM 11Zf(+). Further manipulations of the 5' and 3' ends of these cDNAs have been carried out, and these cDNAs have been further subcloned into other expression vectors such as pGEMEX-1, pET 3b, and the yeast shuttle vectors YEpsec 1 and pEMBLyex 4, and pPIC 9 and pHIL D2. Also provided are host cells transformed with expression vectors containing DNA molecules encoding proteins having Factor C-like enzymatic activity, methods of producing such proteins, methods for purifying Factor C zymogens, and methods for protecting Factor C zymogens from autoactivation by Gram negative bacterial endotoxin while the proenzyme is being purified and/or processed from amoebocyte lysates or from recombinant clones, or during storage or subsequent handling. This protection is afforded by the addition of 5-30% Me.sub.2 SO, which reversibly inhibits the Factor C zymogen.
Source: USPTO / EPO open patent data. Objective bibliographic and citation counts.