PMID- 10648640
OWN - NLM
STAT- MEDLINE
DCOM- 20000302
LR  - 20141120
IS  - 0026-895X (Print)
IS  - 0026-895X (Linking)
VI  - 57
IP  - 2
DP  - 2000 Feb
TI  - Photoaffinity labeling and purification of ZG-16p, a high-affinity 
      dihydropyridine binding protein of rat pancreatic zymogen granule membranes that 
      regulates a K(+)-selective conductance.
PG  - 308-16
AB  - In rat pancreatic zymogen granules (ZG), an ATP-sensitive K(+) conductance and a 
      Cl(-) conductance have been characterized that are inversely regulated by an 
      approximately 65-kDa multidrug resistance P-glycoprotein (mdr1) gene product. In 
      search of a label for purification of this protein, we found that the 
      dihydropyridine derivative (-)-[(3)H]BZDC-DHP, a recently developed high-affinity 
      ligand for Mdr1, binds with similar affinity to ZG membranes (ZGM) (K(d) = 6.2 
      nM). Binding was inhibited by nanomolar concentrations of the L-type Ca(2+) 
      channel blockers azidopine and verapamil and by micromolar concentrations of the 
      K(+) channel blockers glibenclamide and quinidine. Inhibition by glibenclamide 
      was noncompetitive. The Mdr1 modulators cyclosporin A and vinblastine did not 
      inhibit binding, which is different from Mdr1. In addition, only (+/-)-BZDC-DHP, 
      azidopine, and verapamil selectively inhibited the K(+) conductance in ZGs, 
      whereas the Cl(-) conductance was not affected. In photoaffinity labeling 
      experiments, (-)-[(3)H]BZDC-DHP surprisingly specifically and selectively labeled 
      a approximately 19-kDa protein in ZGM with a pharmacological profile identical 
      with the high-affinity binding site but did not label a 65-kDa protein. The 
      19-kDa protein was purified by ion exchange chromatography and SDS-polyacrylamide 
      gel electrophoresis and sequenced. The sequence obtained corresponds to ZG-16p, a 
      recently cloned ZG protein with no apparent homology to Mdr1. The identity of the 
      19-kDa protein was confirmed by immunoprecipitation of (-)-[(3)H]BZDC-DHP-labeled 
      ZGM with an anti-ZG-16p antibody. Furthermore, it is shown that ZG-16p is 
      associated with the ZGM. We propose that ZG-16p, as part of the submembranous 
      granule matrix, regulates the ATP-sensitive K(+) conductance of ZGs.
FAU - Braun, M
AU  - Braun M
AD  - II. Department of Physiology, Medical Faculty, University of Saarland, 
      Homburg/Saar, Germany.
FAU - Thevenod, F
AU  - Thevenod F
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Mol Pharmacol
JT  - Molecular pharmacology
JID - 0035623
RN  - 0 
      (2,6-dimethyl-4-(2-(trifluoromethyl)phenyl)-1,4-dihydropyridine-3,5-dicarboxylic 
      acid (2-(3-(4-benzoylphenyl)propionylamino)ethyl) ester ethyl ester)
RN  - 0 (Affinity Labels)
RN  - 0 (Benzophenones)
RN  - 0 (Dihydropyridines)
RN  - 0 (Enzyme Precursors)
RN  - 0 (Lectins)
RN  - 0 (Photosensitizing Agents)
RN  - 0 (Potassium Channel Blockers)
RN  - 0 (ZG16 protein, rat)
RN  - 7M8K3P6I89 (1,4-dihydropyridine)
RN  - 8L70Q75FXE (Adenosine Triphosphate)
SB  - IM
MH  - Adenosine Triphosphate/metabolism
MH  - Affinity Labels
MH  - Animals
MH  - Benzophenones/pharmacology
MH  - Binding Sites
MH  - Dihydropyridines/metabolism/pharmacology
MH  - Enzyme Precursors/*isolation & purification/metabolism
MH  - In Vitro Techniques
MH  - Lectins/*isolation & purification/metabolism
MH  - Male
MH  - Pancreas/enzymology/*metabolism
MH  - Photosensitizing Agents/pharmacology
MH  - *Potassium Channel Blockers
MH  - Rats
MH  - Rats, Wistar
EDAT- 2000/01/29 09:00
MHDA- 2000/03/04 09:00
CRDT- 2000/01/29 09:00
PHST- 2000/01/29 09:00 [pubmed]
PHST- 2000/03/04 09:00 [medline]
PHST- 2000/01/29 09:00 [entrez]
PST - ppublish
SO  - Mol Pharmacol. 2000 Feb;57(2):308-16.