Characterisation of nuclear pore complex oxalate binding protein from human kidney.
Both rat and human kidney nuclei exhibited time and pH dependent oxalate or histone-oxalate uptake which was inhibited by anion transport inhibitor, 4,4'-dithiocyanostilbene-2,2'-disulphonic acid. Sodium chloride had no effect. Nuclear membrane had oxalate binding at pH 7.4. Extraction of nuclear membrane by Triton-high salt mixture showed maximal oxalate binding activity with nuclear pore complex while nuclear lamin had no oxalate binding. The rat and human kidney nuclear pore complex showed oxalate binding of 144 and 220 pmoles/mg protein respectively. Subsequent purification of the protein on diethyl amino ethyl-Sephadex A 50 column and Sephadex G-200 column yielded 4-fold purification. The protein revealed a molecular weight of 205 kDa on SDS-PAGE. The protein was found to be saturable at 2 microM oxalate and had a Kd of 2.98 pM and a Bmax of 197 pmoles. Antibody for 205 kD was separated from primary biliary cirrhosis serum containing auto antibody against 205 kDa using affinity column chromatography. The oxalate binding activity as well as the nuclear uptake of oxalate or histone-oxalate were inhibited by its antibody.
Selvam
R
R
Department of Medical Biochemistry, Dr A.L.M. Postgraduate Institute of Basic Medical Sciences, University of Madras, Chennai, India.
Vijaya
R
R
Sivakamasundari
P
P
D000818:Animals; D006801:Humans; D008297:Male; D051381:Rats; D001323:Autoantibodies / Q000378:metabolism; D002250:Carbon Radioisotopes; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002846:Chromatography, Affinity; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006657:Histones / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000378:metabolism; D007700:Kinetics; D008105:Liver Cirrhosis, Biliary / Q000097:blood; D022022:Nuclear Pore / Q000378:metabolism; D029543:Nucleocytoplasmic Transport Proteins; D017830:Octoxynol / Q000494:pharmacology; D010070:Oxalates / Q000097:blood; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D011485:Protein Binding; D017208:Rats, Wistar; D012965:Sodium Chloride / Q000494:pharmacology; D013997:Time Factors; D028861:Nuclear Pore Complex Proteins
D001323:Autoantibodies; D002250:Carbon Radioisotopes; D002352:Carrier Proteins; D003902:Detergents; D006657:Histones; D029543:Nucleocytoplasmic Transport Proteins; D017830:Octoxynol; C477940:oxalate binding protein, human; C477939:oxalate binding protein, rat; D010070:Oxalates; D010980:Platelet Membrane Glycoproteins; D012965:Sodium Chloride; C063865:NUP210 protein, human; D028861:Nuclear Pore Complex Proteins