Identification of amino acids that are critical to the processivity function of respiratory syncytial virus M2-1 protein.
The M2-1 protein of respiratory syncytial virus (RSV) is a transcription processivity factor that is essential for virus replication. The function of RSV M2-1 protein can be examined by using an RSVlacZ minigenome assay in vitro since the expression of the lacZ gene is dependent on M2-1. The M2-1 protein of pneumonia virus of mice (PVM), also a member of the Pneumovirus genus, functions poorly in the RSVlacZ minigenome assay despite conservation of the Cys(3)-His(1) motif at its N terminus and an overall 40% amino acid identity with RSV M2-1. To identify the amino acids responsible for the differences between these two proteins, two chimeric proteins were constructed. The RSV/PVM (RP) M2-1 chimera that contains the N-terminal 30 amino acids from RSV and the remaining C-terminal 148 amino acids from PVM maintained a level of activity at an ca. 36% of RSV M2-1. However, the PVM/RSV (PR) M2-1 chimera with the N-terminal 29 amino acids from PVM and 164 amino acids from RSV had an activity of <5% of RSV M2-1, indicating that the functional determinants are mainly located in the N terminus of M2-1. Mutagenesis of the N terminus of PR M2-1 and RSV M2-1 identified that Leu-16 and Asn-17 of RSV M2-1 are critical to the M2-1 function. In addition, several charged residues in the N terminus of RSV M2-1 also contributed to the functional integrity of M2-1.
Zhou
Helen
H
MedImmune Vaccines, Inc., Mountain View, California 94043, USA.
Cheng
Xing
X
Jin
Hong
H
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D016679:Genome, Viral; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D029122:Murine pneumonia virus / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D018113:Respiratory Syncytial Virus, Human / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D014764:Viral Proteins