Proteomic analysis of human O {sup 6}-methylguanine-DNA methyltransferase by affinity chromatography and tandem mass spectrometry

Journal Article · Fri Dec 02 04:00:00 EST 2005 · Biochemical and Biophysical Research Communications

DOI:https://doi.org/10.1016/j.bbrc.2005.09.177· OSTI ID:20713461

Niture, Suryakant K ^[1]; Doneanu, Catalin E ^[2]; Velu, Chinavenmeni S ^[1]; Bailey, Nathan I ^[1]; Srivenugopal, Kalkunte S ^[1]

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1. Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, TX 79106 (United States)
2. Mass Spectrometry Center, Department of Medicinal Chemistry, University of Washington, Seattle, WA 98195 (United States)

Recent evidence suggests that human O {sup 6}-methylguanine-DNA methyltransferase (MGMT), a DNA repair protein that protects the genome against mutagens and accords tumor resistance to many anticancer alkylating agents, may have other roles besides repair. Therefore, we isolated MGMT-interacting proteins from extracts of HT29 human colon cancer cells using affinity chromatography on MGMT-Sepharose. Specific proteins bound to this column were identified by electrospray ionization tandem mass spectrometry and/or Western blotting. These procedures identified >60 MGMT-interacting proteins with diverse functions including those involved in DNA replication and repair (MCM2, PCNA, ORC1, DNA polymerase {delta}, MSH-2, and DNA-dependent protein kinase), cell cycle progression (CDK1, cyclin B, CDK2, CDC7, CDC10, 14-3-3 protein, and p21{sup waf1/cip1}), RNA processing and translation (poly(A)-binding protein, nucleolin, heterogeneous nuclear ribonucleoproteins, A2/B1, and elongation factor-1{alpha}), several histones (H4, H3.4, and H2A.1), and topoisomerase I. The heat shock proteins, HSP-90{alpha} and {beta}, also bound strongly with MGMT. The DNA repair activity of MGMT was greatly enhanced in the presence of interacting proteins or histones. These data, for the first time, suggest that human MGMT is likely to have additional functions, possibly, in sensing and integrating the DNA damage/repair-related signals with replication, cell cycle progression, and genomic stability.

• OSTI ID: 20713461
• Journal Information: Biochemical and Biophysical Research Communications, Journal Name: Biochemical and Biophysical Research Communications Journal Issue: 4 Vol. 337; ISSN 0006-291X; ISSN BBRCA9
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
AFFINITY
ALKYLATION
CELL CYCLE
CHEMOTHERAPY
CHROMATOGRAPHY
DNA DAMAGES
DNA REPAIR
DNA REPLICATION
HEAT-SHOCK PROTEINS
HISTONES
LARGE INTESTINE
MASS SPECTROSCOPY
NEOPLASMS
RNA PROCESSING