The XPA protein is a zinc metalloprotein with an ability to recognize various kinds of DNA damage

Mutat Res. 1994 Nov;315(3):229-37. doi: 10.1016/0921-8777(94)90034-5.

Abstract

The XPA (xeroderma pigmentosum group A) gene encodes a protein of 273 amino acids with a zinc finger motif. The human XPA cDNA was placed in an Escherichia coli expression vector for the synthesis of the recombinant XPA protein. The molecular weight of the wild-type protein was about 40 kDa in SDS-PAGE. Microinjection of the wild-type protein specifically restored the defect of UV-induced unscheduled DNA synthesis in XP-A cells. Thus, the bacterially expressed XPA protein retains biochemical properties identical to those of natural sources. The wild-type protein binds preferentially to UV-, cis-diamminedichloroplatinum(II) (cisplatin)- or osmium tetroxide (OsO4)-damaged DNA as assayed by retention on nitrocellulose filters. In addition, the data from atomic absorption and UV-CD spectra revealed that the wild-type protein is a zinc metalloprotein with secondary structure. Furthermore, the mutant protein, of which the cysteine-103 residue in the zinc finger motif was replaced with serine, has a vastly different protein conformation resulting in a loss of XP-A correcting and DNA-binding activities. These findings indicate that the XPA protein is a zinc-binding protein with affinity for various DNA damages, and a cysteine residue in the C4-type zinc finger motif is indispensable for normal protein conformation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids / analysis
  • Cell Line
  • DNA / biosynthesis
  • DNA / metabolism*
  • DNA Damage*
  • DNA, Viral / radiation effects
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Escherichia coli / genetics
  • Humans
  • Metalloproteins / chemistry
  • Metalloproteins / genetics
  • Metalloproteins / metabolism*
  • Microinjections
  • Molecular Weight
  • Protein Binding
  • Protein Conformation
  • Recombinant Fusion Proteins / biosynthesis
  • Ultraviolet Rays
  • Xeroderma Pigmentosum
  • Xeroderma Pigmentosum Group A Protein
  • Zinc / analysis*

Substances

  • Amino Acids
  • DNA, Viral
  • DNA-Binding Proteins
  • Metalloproteins
  • Recombinant Fusion Proteins
  • XPA protein, human
  • Xeroderma Pigmentosum Group A Protein
  • DNA
  • Zinc