Genetic diversity and response to IFN of the NS3 protease gene from clinical strains of the hepatitis C virus.
The N-terminal one-third of the hepatitis C virus nonstructural gene 3 (NS3) codes for a serine protease. To investigate natural genetic diversity of this enzyme a nested PCR reaction was developed to obtain NS3 protease sequence data directly from patient strains. This data was used to determine genetic diversity, phylogenetic and evolutionary rates, and selection of variants by interferon therapy. The potential effect of genetic diversity on enzyme structure using molecular modeling was also attempted. Results show significant variability in clinical HCV strains at both the nucleotide (30.2% for 1a and 25.8% for 1b) and amino acid sequences (11.0% for 1a and 9.9% for 1b). Phylogenic analysis shows two distinct clades with two HCV isolates grouping as a sister clade to 1b. Structural analysis reveals that most mutations lie in the N-terminus of the enzyme. When strains were sorted as to whether or not the patient had received antiviral therapy, no difference was found in the number or locations of mutations in 1a strains. However, 1b strains demonstrated an overall drop in the number of positions that were mutated. This study demonstrates significant differences among natural strains that may pose a problem for structure based drug development.
Holland-Staley
C A
CA
Infectious Disease Research, Henry Ford Hospital, Detroit, Michigan, USA. chollandstaley@beaumont.edu
Kovari
L C
LC
Golenberg
E M
EM
Pobursky
K J
KJ
Mayers
D L
DL
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000627:therapeutic use*; D004359:Drug Therapy, Combination; D005260:Female; D005814:Genes, Viral; D014644:Genetic Variation; D016174:Hepacivirus / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D006526:Hepatitis C / Q000188:drug therapy / Q000821:virology*; D006801:Humans; D016898:Interferon-alpha / Q000627:therapeutic use*; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012254:Ribavirin / Q000627:therapeutic use; D016415:Sequence Alignment; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D000998:Antiviral Agents; D016898:Interferon-alpha; D012254:Ribavirin; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases