Structural and biophysical analysis of sequence insertions in the Venezuelan Equine Encephalitis Virus macro domain.
Random transposon insertions in viral genomes can be used to reveal genomic regions important for virus replication. We used these genomic data to evaluate at the protein level the effect of such insertions on the Venezuelan Equine Encephalitis Virus nsP3 macro domain. The structural analysis showed that transposon insertions occur mainly in loops connecting the secondary structure elements. Some of the insertions leading to a temperature sensitive viral phenotype (ts) are close to the cleavage site between nsP2 and nsP3 or the ADP-ribose binding site, two important functions of the macro domain. Using four mutants mimicking the transposon insertions, we confirmed that these insertions can affect the macro domain properties without disrupting the overall structure of the protein.
CNRS, AFMB UMR 7257, 13288, Marseille, France; Aix-Marseille Université, AFMB UMR 7257, 13288, Marseille, France.
Jaime
J
Guillén
CNRS, AFMB UMR 7257, 13288, Marseille, France; Aix-Marseille Université, AFMB UMR 7257, 13288, Marseille, France.
Julie
J
Lichière
CNRS, AFMB UMR 7257, 13288, Marseille, France; Aix-Marseille Université, AFMB UMR 7257, 13288, Marseille, France.
Nadia
N
Rabah
Center for Genome Sciences, US Army Medical Research Institute of Infectious Disease, 1425 Porter Street, Fort Detrick, MD 21702, United States.
Brett F
BF
Beitzel
CNRS, AFMB UMR 7257, 13288, Marseille, France; Aix-Marseille Université, AFMB UMR 7257, 13288, Marseille, France.
Bruno
B
Canard
CNRS, AFMB UMR 7257, 13288, Marseille, France; Aix-Marseille Université, AFMB UMR 7257, 13288, Marseille, France. Electronic address: bruno.coutard@afmb.univ-mrs.fr.
Bruno
B
Coutard
D055592:Biophysical Phenomena; D004666:Encephalitis Virus, Venezuelan Equine / Q000235:genetics* / Q000502:physiology*; D008958:Models, Molecular; D016254:Mutagenesis, Insertional; D011487:Protein Conformation; D017361:Viral Nonstructural Proteins / Q000737:chemistry* / Q000235:genetics*; D014779:Virus Replication
D017361:Viral Nonstructural Proteins; C089383:nsp3 protein, alphavirus
Alphavirus; Biophysical studies; Macro domain; Non-structural protein; Sequence insertion; Structure