In order to study the possible mechanism of the cellular proteins involved in the process of replication of HCV RNA. UV cross-linking experiment was used to identify the cellular proteins which bind to the 3′-end of HCV intermediate negative-strand RNA. Results showed that a protein with approximate molecular mass of 45 ku(p45) cross-linked with the 3′-end 131～278 nt of HCV intermediate negative-strand RNA. The p45 presented in various cell lines of different origins. The amounts of RNA-protein complexes increased with increasing amounts of cellular extracts. Non-homologous proteins and RNA transcripts could not compete for the binding between p45 and the 3′-end of HCV intermediate negative-strand RNA. These results suggested that the cellular protein p45 can specifically bind to the secondary structure of the 3′-end of HCV intermediate negative-strand RNA, thus p45 may play an important role in HCV RNA replication.