FIG 5.
Analysis of BHV-1 UL49.5/gM covalently linked complex and gM maturation. Lysates of BHV-1 wt, C42S, VP22Δ, and the double mutant UL49.5 C42S/VP22Δ virus-infected MDBK cells were separated by SDS-PAGE under reducing (+DTT) or nonreducing (−DTT) conditions and immunoblotted with either anti-UL49.5-specific (A) or anti-gM-specific (B) antibodies. Note that under nonreducing conditions (−DTT), the UL49.5 C42S mutant protein expressed by the C42S and double UL49.5 C42S/VP22Δ mutant viruses did not comigrate with gM (A), indicating the loss of the C42-mediated covalent bond between UL49.5 C42S and gM. An approximately 90-kDa band (*1) was detected in all of the virus-infected cell lysates. In addition, an approximately 80-kDa band (*2) was detected in wt- and UL49.5 C42S virus-infected lysates. Note that in the case of the C42S and double UL49.5 C42S/VP22Δ mutant viruses, only the 36-kDa immature form of gM (gM-ER) and its 72-kDa homodimer were detected (B). (C) Immunoblotting of the mock-infected and corresponding virus-infected lysates with anti-gC-specific antibody served as a loading control.