Identification of dengue virus binding proteins using affinity chromatography.
Several studies have identified putative dengue virus receptors using virus overlay protein binding assays (VOPBA) with some apparent success. Given that this technique relies upon the use of electrophoresis of proteins through polyacrylamide gels with varying amounts of protein denaturation, the physiological relevance of the proteins isolated is open to question. To address this issue a Sepharose 4B-dengue virus serotype 2-affinity column was constructed to selectively bind dengue virus binding proteins from HepG2 (liver) cell membrane preparations. Results show that GRP78, but not the 37/67 kDa high affinity laminin receptor, was specifically bound by the column. This result is consistent with earlier work and shows that while affinity chromatography may provide a useful adjunct to VOPBA based studies particularly in cases where proteins maybe sensitive to denaturation, proteins isolated by VOPBA can be physiologically relevant.
Molecular Pathology Laboratory, Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus, 25/25 Phuttamonthol Sai 4, Salaya, Nakorn Pathom 73170, Thailand.
Supranee
S
Upanan
Atichat
A
Kuadkitkan
Duncan R
DR
Smith
D002352:Carrier Proteins / Q000302:isolation & purification*; D002460:Cell Line; D002846:Chromatography, Affinity / Q000379:methods; D003716:Dengue Virus / Q000145:classification* / Q000302:isolation & purification; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins / Q000302:isolation & purification; D006801:Humans; D008099:Liver; D018832:Molecular Chaperones / Q000302:isolation & purification; D012703:Serotyping / Q000379:methods; D014764:Viral Proteins / Q000302:isolation & purification*
D002352:Carrier Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D014764:Viral Proteins