Evidence that a second stereochemical centre in diacylglycerols defines interaction at the recognition site on protein kinase C

Bonser, Robert W.; Thompson, Neil T.; Hodson, H. F.; Beams, Richard M; Garland, L.G.

doi:10.1016/0014-5793(88)80112-1

Cited by 22 publications

(4 citation statements)

References 23 publications

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“…These observations find analogy with studies on the optically active diglycerides; (S)-diolein activates PKC but neither its (R)-enantiomer nor its 1,3-isomer shows substantial activity (5,6). Interestingly, it was reported that 3-methyldiglycerides are active agonists but show unequal activities (23). This suggests that the C29 and C30 centers of DAT correspond to the C2 and C3 centers of diglycerides, respectively.…”

Section: Discussionmentioning

confidence: 99%

Structural basis of protein kinase C activation by tumor promoters.

Nakamura

Kishi

Pajares

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

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Protein kinase C (PKC) is an important enzyme that helps govern cell metabolism and growth. The enzyme is physiologically activated when an (S)-diglyceride binds to its own regulatory domain. The saturable binding site of the regulatory domain can also be bound by any of a group of structurally diverse tumor promoters, including debromoaplysiatoxins (DATs), phorbol esters, ingenols, teleocidins, and bryostatins. The question of how the same binding site can be the target of these structurally diverse molecules is of considerable importance and is addressed in this article. The relatively rigid structure of DAT and the fact that it possesses a diglyceride moiety renders it an ideal starting template. Structure-activity studies with PKC reveal that the C29 but not the C30 stereocenter of DAT is critical for activity. Furthermore, 3-deoxy-DAT and DAT are equipotent as PKC activators, hence the C3 hydroxyl group of DAT is not critical for activity. Straightforward structural considerations show that the C30 hydroxyl group ofDAT matches the C3 hydroxyl group of diglyceride, the C29 stereocenter of DAT matches the C2 stereocenter of (S)-diglyceride, and the C1 ester moiety of DAT matches the C2 ester moiety of diglyceride. Based on these studies and on published structure-activity observations on other tumor promoters, a structural hypothesis is developed to account for the chemical mechanism of tumor promoter action. Experimentally testable predictions are made concerning the interactions with PKC of several classes of tumor PKC activators.A central question in the mechanism of protein kinase C (PKC) action is concerned with the nature of the activation process. This physiologically important enzyme is activated when a diglyceride binds to an effector site on its regulatory domain (1, 2). The diglyceride itself is produced as a consequence of the regulated hydrolysis of phosphatidylinositol phosphates by a specific phospholipase (3). The interaction of PKC with diglyceride is stoichiometric (4) and stereospecific, with the (S)-enantiomer being active (5, 6). The specificity of this process has been further probed with diglyceride analogs (7-9). These studies have shown an unusual degree of specificity directed toward the glycerol backbone and the hydrophilic moieties of the diglycerides. The strict specificity shown toward the diglycerides must be contrasted with the behavior of PKC toward the tumor promoters, including phorbol esters, ingenols, aplysiatoxins, and teleocidins (see Fig. 7). All these structurally diverse molecules share PKC as their common target and appear to function by binding to and activating the enzyme permanently, whereas the diglycerides bind and activate temporarily (10)(11)(12).Given that it is highly likely that the various tumor promoters and active diglycerides bind to the same site on the regulatory domain ofPKC, it must be possible to demonstrate structural commonalities in these various agents. Several models, generated by various computer-driven modeling protocols, have alread...

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Section: Discussionmentioning

confidence: 99%

Structural basis of protein kinase C activation by tumor promoters.

Nakamura

Kishi

Pajares

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

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“…These changes were interpreted as destabilization of bilayers and were proposed to be responsible for the ability of DAGs to activate PK-C and phospholipases. Both the degree of unsaturation and the length of the DAG fatty acid chain affect its capacity to activate PK-C (Kishimoto et al, 1980;Cabot and Jaken, 1984;Lapetina et al, 1985;Bonser et al, 1988). It was also demonstrated that variations in the hydrophobic part of the activating phospholipids affect both the lipid-dependent activation of PK-C and the action of DAG (Snoek et al, 1988).…”

Section: Introductionmentioning

confidence: 99%

Effects of diacylglycerols and Ca2+ on structure of phosphatidylcholine/phosphatidylserine bilayers

Goldberg

Lester

Borchardt

et al. 1994

Biophysical Journal

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The combined effects of the diacylglycerols (DAGs) with the various acyl chains and Ca2+ on the structure of phosphatidylcholine/phosphatidylserine (4:1 mole/mole) bilayers were studied using 2H- and 31P NMR. The following DAG- and Ca(2+)-induced bilayer perturbations were identified. 1) Increased tendency to form nonbilayer lipid phases was induced by diolein or stearoylarachidonoylglycerol, and was synergistically enhanced by the addition of Ca2+. 2) "Transverse" bilayer perturbation was induced by dioctanoylglycerol. The addition of this DAG caused increased ordering of the phospholipid acyl side chains in the region adjacent to the headgroup, with the concomitant decrease of the order toward the bilayer interior. 3) Separation of the phosphatidylcholine and phosphatidylserine bilayer components was induced by combinations of relatively high (1:5 mole/mole to phosphatidylserine) Ca2+ and 25 mol% (to the phospholipids) of diolein, stearoylarachidonoylglycerol, or oleoylacetylglycerol. 4) Lateral phase separation of the bilayers on the regions of different fluidities was induced by dipalmitin. These physicochemical effects were correlated with the effects of these DAGs and Ca2+ on the activity of protein kinase C. The increased tendency to form nonbilayer lipid phases and the transverse bilayer perturbations correlated with the increased protein kinase C activity, whereas the actual presence of the nonbilayer lipid phases, as well as the separation of the phosphatidylcholine and phosphatidylserine components, was associated with the decrease in the protein kinase C activity. The lateral phase separation of the bilayer on gel-like and liquid crystalline regions did not have an effect on the activity of the enzyme. These results demonstrate the importance of the physicochemical properties of the membranes in the process of activation of protein kinase C.

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“…This indicates that the presence of the acidic PS headgroup with all of the cofactors is not sufficient for PK-C activation, which also requires the presence of unsaturated acyl chains on either the PS or phosphatidylchoine (PC) ipid component. Both the degree of unsaturation and the`ngth of the DAG fatty acid chain affect its capacity to activate PK-C (Kishimoto et al, 1980;Cabot and Jaken, 1984;Lapetina et al, 1985;Bonser et al, 1988;Snoek et al, 1988;Goldberg et al, 1994). These studies indicate that DAGs generate physical perturbations in the biiayer structure that may be sensed by PK-C and modulate its activation.…”

Section: Introductionmentioning

confidence: 99%

Effects of diacylglycerols on conformation of phosphatidylcholine headgroups in phosphatidylcholine/phosphatidylserine bilayers

Goldberg

Lester

Borchardt

et al. 1995

Biophysical Journal

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The effects of five diacylglycerols (DAGs), diolein, 1-stearoyl,2-arachidonoyl-sn-glycerol, dioctanoylglycerol, 1-oleoyl,2-sn-acetylglycerol, and dipalmitin (DP), on the structure of lipid bilayers composed of mixtures of phosphatidylcholine and phosphatidylserine (4:1 mol/mol) were examined by 2H nuclear magnetic resonance (NMR). Dipalmitoylphosphatidylcholine deuterated at the alpha- and beta-positions of the choline moiety was used to probe the surface region of the membranes. Addition of each DAG except DP caused a continuous decrease in the beta-deuteron quadrupole splittings and a concomitant increase in the alpha-deuteron splittings indicating that DAGs induce a conformational change in the phosphatidylcholine headgroup. Additional evidence of conformational change was found at high DAG concentrations (> or = 20 mol%) where the alpha-deuteron peaks became doublets indicating that the two alpha-deuterons were not equivalent. The changes induced by DP were consistent with the lateral phase separation of the bilayers into gel-like and fluid-like domains with the phosphatidylcholine headgroups in the latter phase being virtually unaffected by DP. The DAG-induced changes in alpha-deuteron splittings were found to correlate with DAG-enhanced protein kinase C (PK-C) activity, suggesting that the DAG-induced conformational changes of the phosphatidylcholine headgroups are either directly or indirectly related to a mechanism of PK-C activation. 2H NMR relaxation measurements showed significant increase of the spin-lattice relaxation times for the region of the phosphatidylcholine headgroups, induced by all DAGs except DP. However, this effect of DAGs did not correlate with the DAG-induced activation of PK-C.

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Evidence that a second stereochemical centre in diacylglycerols defines interaction at the recognition site on protein kinase C

Cited by 22 publications

References 23 publications

Structural basis of protein kinase C activation by tumor promoters.

Structural basis of protein kinase C activation by tumor promoters.

Effects of diacylglycerols and Ca2+ on structure of phosphatidylcholine/phosphatidylserine bilayers

Effects of diacylglycerols on conformation of phosphatidylcholine headgroups in phosphatidylcholine/phosphatidylserine bilayers

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