|Budget Amount *help
¥3,300,000 (Direct Cost : ¥3,300,000)
Fiscal Year 1999 : ¥900,000 (Direct Cost : ¥900,000)
Fiscal Year 1998 : ¥2,400,000 (Direct Cost : ¥2,400,000)
The RNA-dependent NA polymerase of influenza A virus consists of three subunits, pB1, PB2 and PA. PB1 is a catalytic subunit of NA synthesis. PB2 confers a transcriptase activity to PB1. PA provides replicase activity and performs a recently identified unique protease activity. In order to map its functional domains, we generated site-specific antibodies. RNA synthesis activity was mapped in amino acids 1-159 of PB2 and 501-617 of PA. Cap-I dependent RNase activity was mapped in amino acids 1-159 and 305-559 of PB2, 1-222 of PB1, and 601-716 of PA. Cap-binding site was mapped in amino acids 402-559 of PB2.
We previously determined that the C-terminal 158 amino acids of PB1 bound to the N-termina1 249 amino acid stretch of PB2, while the N-terminal 140 amino acids ofPB1 bound to the C-terminal two third region of PA. This time, we have narrowed the PB1-PB2 binding site in 28 residues of PB2 and 180 residues of PB1, and the PB1-PA binding site in 25 residues of PB1 and about 200 residues of PA.
In order to expand our knowledge into other viral RNA-dependent RNA polymerases, we established in vitro transcription system of hepatitis C RNA polymerase.