title: Philibertain g I, the most basic cysteine endopeptidase purified from the latex of Philibertia gilliesii Hook. et Arn. (Apocynaceae).
abstract: A new papain-like cysteine peptidase isolated from latex of Philibertia gilliesii Hook. et Arn., Apocynaceae (formerly Asclepiadaceae) has been purified and characterized. The enzyme, named philibertain g I, is the most basic component present in latex extracts and was purified by acetone fractionation followed by cation exchange chromatography (SP-Sepharose HR) using FPLC system. Homogeneity was confirmed by SDS-PAGE and mass spectroscopy (MS). Molecular mass of the enzyme was 23,530 Da (MALDI-TOF MS), its isoelectric point was >10.25, and maximum proteolytic activity (casein) was achieved at pH 7-8. The new protease was inhibited by E-64 a cysteine peptidases inhibitor. Km was 0.15 mM, using PFLNA as substrate. The N-terminal sequence of philibertain g I (LPASVDWRKEGAVLPIRHQGQCG) was compared with those of twenty plant proteases. Philibertain g I showed the higher degree of identity (73%) with caricain, one of the Carica papaya endopepetidases.
authors.affiliation: CENPAT-CONICET Centro Nacional Patagónico, Blvd. Brown s/n, 9120, Puerto Madryn, Chubut, Argentina.
authors.forename: C
authors.initials: C
authors.lastname: Sequeiros
authors.forename: M J
authors.initials: MJ
authors.lastname: Torres
authors.forename: S A
authors.initials: SA
authors.lastname: Trejo
authors.forename: J L
authors.initials: JL
authors.lastname: Esteves
authors.forename: C L
authors.initials: CL
authors.lastname: Natalucci
authors.forename: L M I
authors.initials: LM
authors.lastname: López
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D029062:Apocynaceae / Q000737:chemistry / Q000201:enzymology; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D007840:Latex / Q000737:chemistry*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D016415:Sequence Alignment
chemical_list: D007840:Latex; D010940:Plant Proteins; D003546:Cysteine Endopeptidases