Role of variable regions A and B in receptor binding domain of amphotropic murine leukemia virus envelope protein.
For the amphotropic murine leukemia virus (MuLV), a 208-amino-acid amino-terminal fragment of the surface unit (SU) of the envelope glycoprotein is sufficient to bind to its receptor, Pit2. Within this binding domain, two hypervariable regions, VRA and VRB, have been proposed to be important for receptor recognition. In order to specifically locate residues that are important for the interaction with Pit2, we generated a number of site-specific mutations in both VRA and VRB and analyzed the resulting envelope proteins when expressed on retroviral vectors. Concurrently, we substituted portions of the amphotropic SU with homologous regions from the polytropic MuLV envelope protein. The amphotropic SU was unaffected by most of the point mutations we introduced. In addition, the deletion of eight residues in a region of VRA that was previously suggested to be essential for Pit2 utilization only decreased titer on NIH 3T3 cells by 1 order of magnitude. Although the replacement of the amino-terminal two-thirds of VRA with the polytropic sequence abolished receptor binding, smaller nonoverlapping substitutions did not affect the function of the protein. We were not able to identify a single critical receptor contact point within VRA, and we suggest that the amphotropic receptor binding domain probably makes multiple contacts with the receptor and that the loss of some of these contacts can be tolerated.
Gene Therapy Laboratories, Norris Cancer Center and Department of Biochemistry and Molecular Biology, University of Southern California School of Medicine, Los Angeles, California 90033, USA.
J Y
JY
Han
Y
Y
Zhao
W F
WF
Anderson
P M
PM
Cannon
D016475:3T3 Cells; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D014644:Genetic Variation; D009052:Leukemia Virus, Murine / Q000235:genetics* / Q000502:physiology*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D011991:Receptors, Virus / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*
D011991:Receptors, Virus; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins