| Abstrakt: |
Although the product of the UL12 gene of herpes simplex virus type 1 (HSV-1) has been shown to possess both exonuclease and endonuclease activitiesin vitro,and deletion of most of the gene within the viral genome results in inefficient production and maturation of infectious virions, the function of the deoxyribonuclease (DNase) activity per se in virus replication remains unclear. In order to correlate thein vitroandin vivoactivities of the protein encoded by UL12, mutant proteins were tested for nuclease activityin vitroby a novel hypersensitivity cleavage assay and for their ability to complement the replication of a DNase null mutant, AN-1. Rabbit reticulocyte lysates programmed with wild-type UL12 RNA cleaved at the same sites cleaved by purified HSV-1 DNase, but distinct from those cleaved by DNase I or micrococcal nuclease. All mutants which lacked DNase activityin vitroalso failed to complement the replication of AN-1 in nonpermissive cells. Likewise, all mutants which contained HSV-1 DNase activity, as detected by the hypersensitivity cleavage assay, were capable of complementing the replication of the DNase null mutant, though to varying extents. Of particular note was the d1-126 mutant protein, which, despite having the same specific activity as the wild-type enzymein vitro,complemented the replication of AN-1 significantly less than the wild-type protein. The results suggest that DNase activity per se is required for efficient replication of HSV-1in vivo.However, residues, including the N-terminal 126 amino acids, which are dispensable for enzymatic activityin vitromay facilitate the accessibility or activity of the proteinin vivo. |
Full Text from ScienceDirect