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Article Dans Une Revue Biochemistry Année : 2001

Peptidic Determinants and Structural Model of Human NDP Kinase B (Nm23-H2) Bound to Single-Stranded DNA

Résumé

Isoform B of human NDP kinase (NDPK-B) was previously identified as a transcription factor stimulating in vitro and ex vivo the transcription of the c-myc oncogene, which involves this enzyme in carcinogenesis. We have studied the enzymatic properties of NDPK-B in the presence of several single-stranded oligonucleotides. We show that the oligonucleotides are competitive inhibitors of the catalytic activity, indicating that the active site acts as a binding template for the anchorage of the oligonucleotide. Furthermore, the presence of a guanine at the 3'-end of several different aptamers increases its affinity 10-fold. To define the surface of the protein contacting the DNA within the nucleoprotein complex, we used single nanosecond laser pulses as the cross-linking reagent and MALDI-TOF mass spectrometry to identify cross-linked peptides purified from proteolytic digests of the cross-linked complex. Using 11-mer and 30-mer single-stranded oligonucleotides, the same three different nucleopeptides were identified after irradiation of the complexes, indicating a common binding mode for these two aptamers. Taken together, these results allowed us to propose a structural model of NDPK-B bound to single-stranded DNA.

Domaines

Chimie

Dates et versions

hal-01636976 , version 1 (17-11-2017)

Identifiants

Citer

Sharona Raveh, J. Vinh, J. Rossier, F. Agou, Michel Véron. Peptidic Determinants and Structural Model of Human NDP Kinase B (Nm23-H2) Bound to Single-Stranded DNA. Biochemistry, 2001, 40 (20), pp.5882-5893. ⟨10.1021/bi001085j⟩. ⟨hal-01636976⟩
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