Table 1.

Site-directed mutagenesis on fidelity checkpoint residues 64, 239, and 241

Amino acid 64Amino acid 239Amino acid 241
Desired variantStatus*New mutationDesired variantStatus*New mutationDesired variantStatus*New mutation
ArgNonviableGlnNonviableAlaNonviable
AsnNonviablePheNonviableArgNonviable
GluNonviableProNonviableAsnNonviable
IsoNonviableTrpNonviableAspNonviable
LeuNonviableTyrNonviableCysNonviable
ProNonviableArgUnstableR239GGlnNonviable
ValNonviableGluUnstableE239GGluNonviable
AspReversionP48K or S164PCysUnstableC239GGlyNonviable
CysReversionS164PAsnUnstableN239SHisNonviable
HisReversionS164PAspReversionLysNonviable
LysReversionS164PThrReversionMetNonviable
MetReversionS164PValReversionPheNonviable
PheReversionA239GHisReversionS299TProNonviable
ThrReversionS164PIsoReversionS299TSerNonviable
TrpReversionS164PLysReversionS299TThrNonviable
TyrReversionP48KMetReversionS299TTrpNonviable
AlaStablePheReversionS299TTyrNonviable
GlnStableGlyStableValNonviable
SerStableSerStableIsoStable
GlyWTAlaWTLeuWT
  • Dashes indicate that no additional, compensatory mutations were identified.

  • *The status indicates whether the mutation was stable over three passages (dark gray), reverted to WT (reversions, lighter gray), changed to another amino acid (unstable, lightest gray), or did not yield viable virus. The wild-type (WT) amino acid at each position is indicated at the bottom of the variant list.

  • New mutation is acquired by unstable or reverted viruses within the first cycle of replication, and all four independent clones gave rise to the same mutation. For G64D, two clones gave P48K, and two clones gave S164P.