Open AccessLarge‐scale purification of hnRNP proteins from HeLa cells by affinity chromatography on ssDNA‐cellulose
Author(s) -
PANDOLFO Massimo,
VALENTINI Omar,
BIAMONTI Giuseppe,
ROSSI Paola,
RIVA Silvano
Publication year - 1987
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1987.tb10563.x
Subject(s) - hela , protease , affinity chromatography , biochemistry , molecular mass , rna , dna , chemistry , microbiology and biotechnology , biology , cell , enzyme , gene
A purification procedure for proteins which bind heterogeneous nuclear RNA (hnRNP proteins) is described. The procedure, which entails standard chromatographic fractionations (single‐stranded DNA cellulose, hydroxyapatite) and detection with specific antibodies, allows a large‐scale preparation of these proteins and the partial separation of different polypeptides. By this method, polypeptides of higher molecular mass (53–55 kDa) can be purified, which are structurally and antigenically related to the ‘canonical’ hnRNP core proteins (34–43 kDa) that constitute the 40S hnRNP complexes. We also show that HeLa cells contain a protease that cleaves hnRNP core proteins to discrete smaller polypeptides of 22–28 kDa. Such protease, which has been partially purified, appears to copurify extensively with some of the hnRNP proteins.