Related Experiment Videos
Identification of a subdomain in the Moloney murine leukemia virus envelope protein involved in receptor binding
A J MacKrell1, N W Soong, C M Curtis
1Gene therapy Laboratories, Norris Cancer Cancer, University of Southern California School of Medicine, Los Angeles 90033, USA.
Abstract:
We have mutated amino acids within the receptor-binding domain of Moloney murine leukemia virus envelope in order to identify residues involved in receptor binding. Analysis of mutations in the region of amino acids 81 to 88 indicates that this region is important for specific envelope-receptor interactions. None of the aspartate 84 (D-84) mutants studied bind measurably, although they are efficiently incorporated into particles. D-84 mutants have titers that correspond to the severity of the substitution. This observation suggests that D-84 may provide a direct receptor contact. Mutations in the other charged amino acids in this domain (R-83, E-86, and E-87) yield titers similar to those of wild-type envelope, but the affinity of the mutant envelope in the binding assay is decreased by nonconservative substitutions in parallel to the severity of the change. These other amino acids may either provide secondary receptor contacts or assist in maintaining a structure in the domain that favors efficient binding. We also studied other regions of high hydrophilicity. Our initial characterization indicates that amino acids 106 to 111 and 170 to 188 do not play a major role in receptor binding. Measurements of relative binding affinity and titer indicate that most mutations in the region of amino acids 120 to 131 did not significantly affect receptor binding. However, SU encoded by mutants H123V, R124L, and C131A as well as C81A could not be detected in particles and therefore did not bind measurably. Therefore, the region encompassed by amino acids 81 to 88 appears to be directly involved in receptor binding.
Insights
Researchers mutated Moloney murine leukemia virus envelope amino acids to pinpoint receptor binding sites. The 81-88 amino acid region, particularly aspartate 84, is crucial for specific viral envelope-receptor interactions.
Area of Science:
- Virology
- Molecular Biology
- Structural Biology
Background:
- Moloney murine leukemia virus (MMLV) utilizes its envelope protein to bind specific cellular receptors, initiating infection.
- Understanding these interactions is key to developing antiviral strategies and comprehending viral entry mechanisms.
Purpose of the Study:
- To identify specific amino acid residues within the MMLV envelope's receptor-binding domain critical for receptor interaction.
- To elucidate the roles of different amino acid regions in mediating envelope-receptor binding affinity and viral infectivity.
Main Methods:
- Site-directed mutagenesis was employed to alter amino acids within the MMLV envelope's receptor-binding domain.
- Mutant envelopes were analyzed for particle incorporation, binding affinity to cellular receptors, and viral titer (infectivity).
Main Results:
- Mutations in the amino acid region 81-88 significantly impacted receptor binding, with D-84 mutants showing no measurable binding.
- While other charged residues (R-83, E-86, E-87) mutations decreased binding affinity, they retained infectivity, suggesting secondary roles or structural support.
- Mutations in regions 106-111, 170-188, and most of 120-131 did not significantly affect receptor binding.
Conclusions:
- The amino acid region 81-88, particularly D-84, is directly involved in specific Moloney murine leukemia virus envelope-receptor binding.
- Specific residues may act as direct contact points or contribute to the structural integrity required for efficient receptor engagement.