A novel multiple affinity purification tag and its use in identification of proteins associated with a cyclin-CDK complex | Zendy

Shwe Sin Honey | Zendy

Open Access

A novel multiple affinity purification tag and its use in identification of proteins associated with a cyclin-CDK complex

Author(s) -

Shwe Sin Honey

Publication year - 2001

Publication title -

nucleic acids research

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 9.008

H-Index - 537

eISSN - 1362-4954

pISSN - 0305-1048

DOI - 10.1093/nar/29.4.e24

Subject(s) - biology , tandem affinity purification , affinity chromatography , biochemistry , cyclin dependent kinase , microbiology and biotechnology , calmodulin binding proteins , cyclin dependent kinase 1 , protein purification , calmodulin , enzyme , cell cycle , cell

A novel multiple affinity purification (MAFT) or tandem affinity purification (TAP) tag has been constructed. It consists of the calmodulin binding peptide, six histidine residues, and three copies of the hemagglutinin epitope. This 'CHH' MAFT tag allows two or three consecutive purification steps, giving high purity. Active Clb2-Cdc28 kinase complex was purified from yeast cells after inserting the CHH tag into Clb2. Associated proteins were identified using mass spectrometry. These included the known associated proteins Cdc28, Sic1 and Cks1. Several other proteins were found including the 70 kDa chaperone, Ssa1.

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