title: Purification, partial characterization and molecular cloning of the novel antiviral protein RC28.
abstract: An antiviral protein, RC28, with anti-herpesvirus activity was purified from a PBS extract of Rozites caperata (Cortinarious caperata) by acetone precipitation followed by gel filtration and ion exchange chromatography. The molecular weight of this protein was 28.251kDa as measured by MALDI-TOF mass spectrography. Our preparation of RC28 inhibited HSV-1 replication in vitro with an IC(50) value of 0.078mg/ml and a therapeutic index >32. The first 30 amino acid residues of RC28 were sequenced by Edman degradation to be MLTYRGKLNWYNYAVNEGFTLILPGXELKV. Based on that sequence, two degenerate primers were designed, the RC28 cDNA fragment was cloned by 3'-RACE, and the rest of the amino acid sequence was inferred from the cDNA sequence. The full-length peptide chain of RC28 has 235 amino acid residues, and was modified after its translation naturally. A search of the literature showed that this sequence has not been reported before and does not belong to any known protein family. A preliminary expression system was also constructed by inserting the cDNA into the PQE-30 vector, and transformed into Escherichia coli.
authors.affiliation: Ministry of Health, West China Hospital, Sichuan University, Chengdu, China.
authors.forename: Meng
authors.initials: M
authors.lastname: Gong
authors.forename: Frank
authors.initials: F
authors.lastname: Piraino
authors.forename: Naihong
authors.initials: N
authors.lastname: Yan
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.forename: Minjie
authors.initials: M
authors.lastname: Xia
authors.forename: Jia
authors.initials: J
authors.lastname: Ma
authors.forename: Jingqiu
authors.initials: J
authors.lastname: Cheng
authors.forename: Xuyang
authors.initials: X
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D001483:Base Sequence; D001487:Basidiomycota / Q000737:chemistry; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D018259:Herpesvirus 1, Human / Q000187:drug effects; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000998:Antiviral Agents; D017931:DNA Primers; D005656:Fungal Proteins; C542559:RC28 protein, Rozites caperata; D011994:Recombinant Proteins