Open AccessRibonuclease U2: cloning, production in Pichia pastoris and affinity chromatography purification of the active recombinant protein
Author(s) -
MartínezRuiz Antonio,
GarcíaOrtega Lucía,
Kao Richard,
Oñaderra Mercedes,
Mancheño José M.,
Davies Julian,
Martínez del Pozo Álvaro,
Gavilanes José G.
Publication year - 2000
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2000.tb09224.x
Subject(s) - pichia pastoris , recombinant dna , ribonuclease , affinity chromatography , rnase p , biochemistry , pichia , complementary dna , endoribonuclease , amino acid , biology , yeast , signal peptide , microbiology and biotechnology , flag tag , enzyme , rna , gene , fusion protein
RNase U2 is an endoribonuclease secreted by the fungus Ustilago sphaerogena . Its genomic DNA ( rnu2 ), containing an intron of 116 bp, has been isolated and cloned. The corresponding cDNA has also been synthesized. The recombinant RNase U2 was successfully produced in Pichia pastoris , fused to the yeast alkaline phosphatase signal peptide. The recombinant RNase U2, purified by affinity chromatography, contains three extra amino acids at its amino‐terminal end and retains the enzymatic and spectroscopic properties of the natural fungal protein.