The effects of hydrostatic pressure on pertussis toxin-catalyzed ribosylation of guanine nucleotide-binding proteins from two congeneric marine fish

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The effects of pressure on pertussis toxin (PTX)-catalyzed [32P]ADP-ribosylation of α-subunits of the guanine nucleotide-binding proteins, Gi and Go, were examined in the presence of the guanyl nucleotides GDP and guanosine 5′-O-[γ-thio]triphosphate (GTPγS) in brain membrane preparations from two congeneric marine fish that live at different depths. In the presence of 100 μM GDP, pressures up to 340 atm had no effect on PTX-catalyzed [32P]ADP-ribosylation in the deeper-occurring species, Sebastolobus altivelis. [32P]ADP-ribosylation was suppressed 37% by 68 atm pressure in the shallower-living S. alascanus. Pressure in the range 204-476 atm inhibited the reaction approximately 54%. In the presence of 100 μM GTPγS, the effects of pressure were identical in the two species. Pressures of 68-340 atm inhibited [32P]ADP-ribosylation approximately 30% in both species; 476 atm inhibited ribosylation 54%. In the shallower-living species, pressure may increase the fraction of G proteins coupled to unoccupied receptors or decrease the efficacy of GDP in promoting the heterotrimeric aggregation state. Pressure appears to enhance the efficacy of GTPγS in dissociating the heterotrimeric holoprotein in both species. The structural similarities of the Gi/Go α-subunits of the two species were compared by mapping partial proteolytic digests of brain homogenates labeled with [32P]ADP by PTX on sodium dodecyl sulfate-polyacrylamide gels. The proteases used were TPCK-treated trypsin, subtilisin Carlsberg and Staphylococcus aureus strain V8 endoproteinase Glu-C. The [32P]ADP-labeled peptide maps of the two species were indistinguishable. The differences in pressure sensitivity between the two species may result from small changes in primary structure and/or post-translational modification of the heterotrimeric subunits. © 1994.

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Comparative Biochemistry and Physiology -- Part B: Biochemistry and

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