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J Baudier

Publications and source records attributed to J Baudier.

At least 37 records · Page 2Linked to original sources
PubMed1992

Phosphorylation of the MARCKS protein (P87), a major protein kinase C substrate, is not an obligatory step in the mitogenic signaling pathway of basic fibroblast growth factor in rat oligodendrocytes.

Basic fibroblast growth factor (bFGF) is a well-characterized peptide hormone that has mitogenic activity for various cell types and elicits a characteristic set of responses on the cell types investigated. In this report we confirmed that bFGF is a potent mitogen for rat brain-derived oligodendrocyte (OL) precursor cells as well as for differentiated OL in secondary culture. bFGF was shown to induce expression of the protooncogene c-fos in OL. The role of protein kinase C (PKC) in mediating bFGF-stimulated proliferation as well as c-fos expression in OL was investigated. The PKC activator phorbol 12-myristate 13-acetate (PMA) stimulated c-fos expression but did not trigger cell proliferation. When PKC was down-regulated by pretreatment of OL with PMA for 20 h, the bFGF-mediated stimulations of OL proliferation and c-fos mRNA expression were still observed, whereas the induction of c-fos mRNA by PMA was totally inhibited. These data demonstrate that the bFGF mitogenic signaling pathway in OLs does not require PKC. On the other hand, bFGF was found to stimulate specifically the phosphorylation of a limited number of PKC substrates in oligodendroglial cells, including the MARCKS protein. The bFGF-dependent phosphorylation of MARCKS protein was totally inhibited when PKC was first down-regulated, indicating that the phosphorylation of this protein is PKC dependent. Tryptic digestion of the phosphorylated MARCKS protein revealed that bFGF stimulated specifically the phosphorylation of the MARCKS protein on a single phosphopeptide. We provide evidence that bFGF also stimulated fatty acylation of the MARCKS protein, which might explain the observed specific bFGF-dependent phosphorylation of this protein in OL. We propose that bFGF-dependent fatty acylation and phosphorylation of the MARCKS protein are not essential for the transduction of the bFGF mitogenic signal but are probably linked to differentiation processes elicited by bFGF on OL.

Animals↗
PubMed1991

A rapid purification method for neurogranin, a brain specific calmodulin-binding protein kinase C substrate.

A rapid purification method is reported for bovine brain neurogranin, a calmodulin-binding protein kinase C (PKC) substrate. This method takes advantage of the fact that the protein remains soluble in 2.5% perchloric acid (PCA) and that it binds to a calmodulin-Sepharose column in the absence of calcium: Other PKC substrate proteins that remain to be identified were also found to share these two properties, suggesting that a class of calmodulin-binding PKC substrates may exist in the brain.

Amino Acid Sequence↗
PubMed1991

The interactions of the brain-specific calmodulin-binding protein kinase C substrate, neuromodulin (GAP 43), with membrane phospholipids.

The interaction of the brain-specific calmodulin-binding protein kinase C (PKC) substrate, neuromodulin (GAP 43), with membrane phospholipids was studied. Specific binding of neuromodulin to negatively charged phospholipids through electrostatic interactions was demonstrated. Comparison of the binding of neuromodulin to acidic phospholipids with that of neurogranin, a newly characterized calmodulin-binding PKC substrate (Baudier J., Deloulme, J. C., Van Dorsselaer, A., Black, D., and Mathes H. (1991) J. Biol. Chem. 266, 229-237) suggested that the conserved basic amino acid sequence which characterizes the two proteins and which corresponds to the PKC phosphorylation and calmodulin binding domain also serves as phospholipid binding site. In the absence of calmodulin, binding of neuromodulin to phosphatidylserine at low concentration parallels its phosphorylation by PKC, suggesting that formation of a ternary complex between neuromodulin, phosphatidylserine, and PKC is required for optimum neuromodulin phosphorylation. In the presence of calmodulin, the binding of neuromodulin to phosphatidylserine is inhibited, resulting in total inhibition of neuromodulin phosphorylation. Our results suggest that, in vivo, phosphorylation of neuromodulin may not only depend on protein kinase C (PKC)1 activation but also on the accessibility of the neuromodulin phosphorylation domain to activated membrane-bound PKC that could regulated by CaM.

Amino Acid Sequence↗
PubMed1991

Purification and characterization of a brain-specific protein kinase C substrate, neurogranin (p17). Identification of a consensus amino acid sequence between neurogranin and neuromodulin (GAP43) that corresponds to the protein kinase C phosphorylation site and the calmodulin-binding domain.

Neurogranin, formerly designated p17 (Baudier, J., Bronner, C., Kligman, D., and Cole, R. D.) (1989) J. Biol. Chem. 264, 1824-1828), a brain-specific in vitro substrate for protein kinase C (PKC), has been purified to homogeneity from bovine forebrain. The purified protein has a molecular mass of 7837.1 +/- 0.5 Da, determined by electrospray mass spectrometry. In the absence of reducing agent, dimers and higher oligomers accumulated. On sodium dodecyl sulfate-polyacrylamide gels the protein monomer migrated abnormally with an apparent molecular mass of 15,000-19,000 Da, depending on the percentage of polyacrylamide. The native protein is blocked at its amino terminus. The majority of the primary amino acid sequence was determined following proteolytic and chemical fragmentation. A comparison of the amino acid sequence of neurogranin with that of the brain-specific PKC substrate neuromodulin, revealed a strikingly conserved amino acid sequence AA(X)KIQA-SFRGH(X)(X)RKK(X)K. The two proteins are not related over the rest of their sequences. Neurogranin was shown to be phosphorylated in hippocampal slices incubated with 32Pi and phorbol esters stimulated neurogranin phosphorylation, suggesting that neurogranin is likely to be an in vivo substrate for PKC. In vitro phosphorylation of neurogranin by PKC produced a shift of the isoelectric point of the protein (pI 5.6) to a more acidic value (pI 5.4). Tryptic digestion of the phosphorylated protein yielded a single phosphopeptide having the sequence IQASFR, where the serine residue is the phosphorylated amino acid. This phosphopeptide is part of the conserved sequence shared with neuromodulin and also corresponds to the PKC phosphorylation site on neuromodulin (Apel, E. D., Byford, M. F., Au, D., Walsh, K. A., and Storm, D. R. (1990) Biochemistry 29, 2330-2335). Evidence was obtained suggesting that neurogranin binds to calmodulin in the absence of Ca2+, a feature that also characterizes neuromodulin. We propose that the amino acid sequence shared by neurogranin and neuromodulin reflects a functional relationship between these two proteins and that the consensus sequence represents a conserved PKC phosphorylation site and a calmodulin binding domain that characterizes a class of brain-specific PKC substrates.

Amino Acid Sequence↗
PubMed1991

Establishment of pure neuronal cultures from fetal rat spinal cord and proliferation of the neuronal precursor cells in the presence of fibroblast growth factor.

A primary culture system of nearly pure neuronal cells from 14-day-old fetal rat spinal cord has been developed by combining a preplating step, the use of a chemically defined serum-free medium, and borated polylysine-coated dishes that prevented the formation of cell aggregates. About 98% of the cells were found to be immunostained with neuron-specific enolase antibodies, confirming their neuronal nature. The cultures are composed essentially of a population of non-motoneurons and contain few motoneurons, characterized by their large size and multipolar aspect, the presence of acetylcholinesterase (AChE), and the intense immunoreaction for growth-associated protein GAP-43. Neuronal precursor cells are also present in these cultures and proliferate during the first 3 days. The addition of bovine brain basic fibroblast growth factor (bFGF) stimulates their proliferation over a period of 2 days, as determined by measurement of [125I]iododeoxyuridine incorporation and by immunocytochemical reaction after bromodeoxyuridine incorporation into nuclei. The proliferating cells were characterized as neurons by immunostaining against neuron-specific enolase. Recombinant human bFGF and bovine brain acidic FGF (aFGF) exerted similar effects. Other growth factors, including epidermal growth factor (EGF), transforming growth factor beta 1 (TGF-beta 1), and thrombin, were without effect on the proliferative activity of these neuronal cells. bFGF has no effect on the survival of motoneurons and on the fiber outgrowth of the whole neuronal population. However, bFGF affects the development of bipolar AChE-positive neurons, probably belonging to the non-motoneuron population. The data indicate that bFGF and aFGF are mitogens for neuroblasts from rat spinal cord in culture and that bFGF influences the development of a subpopulation of spinal neurons that are AChE-positive.

Acetylcholinesterase↗
PubMed1991

Membrane-related oxysterol function: preliminary results on the modification of protein kinase C activity and substrate phosphorylation by 7 beta,25-dihydroxycholesterol.

Oxysterols exhibit a wide variety of biological activities, including potent immunosuppressive effects. 7 beta,25-Dihydroxycholesterol (7,25-OHC), a synthetic oxysterol, has been shown to strongly inhibit the lymphocyte response to different stimuli. This compound has been chosen as a model compound to investigate the mechanisms underlying the immunosuppressive effects of oxysterols. As protein kinase C (PKC) constitutes a key enzyme in the pathways leading to cell activation, we have studied the effect of 7,25-OHC on PKC activity in the cytosolic and particulate fractions of spleen cells. Lymphocytes treated with 7,25-OHC showed a decrease of the relative PKC activity in the particulate fractions compared to control cells. These results are confirmed by the observation that 7,25-OHC also reduces the phosphorylation of the endogenous PKC substrates. Thus oxysterols interfere with two membrane related phenomena, ie the modification of membrane PKC activity and the inhibition of the phosphorylation of the substrates of PKC located in the membrane. Previous results obtained by fluorescence polarisation revealed a modification of the membrane fluidity after oxysterol treatment. Furthermore, it has been demonstrated that oxysterols are incorporated into cell membranes. The alteration of the cell membrane could impair the signal transduction and may explain the immunosuppressive activity of oxysterol. Thus, along with other biological effects previously reported, oxysterols decrease membrane associated PKC activity in immune cells.

Animals↗
PubMed1991

Patterns of aberrant sprouting in Alzheimer's disease.

Alzheimer's disease (AD) is characterized by extensive synaptic and neuronal loss and by plaque formation in the cortex, but the mechanisms responsible for synaptic plasticity in the neocortex are still not completely understood. To analyze the sprouting response in AD cortex, we compared the patterns of GAP-43 with synaptophysin immunoreactivity. In AD, GAP-43 immunohistochemistry revealed extensive sprouting in the hippocampal molecular layer, stratum polymorphous, CA1 region, and prosubiculum. These regions presented abundant anti-GAP-43-immunoreactive coiled fibers and dystrophic neurites in association with plaques. Some of these sprouting structures were colocalized with anti-synapto-physin- and anti-neurofilament-positive neurites. The AD neocortex was characterized by an overall decrease in GAP-43 immunoreactivity accompanied by sprouting neurites in the areas of synaptic pathology. We conclude that GAP-43 might be involved in the mechanisms of synaptic plasticity in the AD cortex, as well as in the process of aberrant sprouting in the neuritic plaques.

Aged↗
PubMed1990

Neuromodulin (GAP43): a neuronal protein kinase C substrate is also present in 0-2A glial cell lineage. Characterization of neuromodulin in secondary cultures of oligodendrocytes and comparison with the neuronal antigen.

Neuromodulin (also called GAP43, G50, F1, pp46), a neural-specific calmodulin binding protein, is a major protein kinase C substrate found in developing and regenerating neurons. Here, we report the immunocytochemical characterization of neuromodulin in cultured 0-2A bipotential glial precursor cells obtained from newborn rat brain. Neuromodulin is also present in oligodendrocytes and type 2 astrocytes (stellate-shaped astrocytes), which are both derived from the bipotential glial 0-2A progenitor cells, but is absent of type 1 astrocytes (flat protoplasmic astrocytes). These results support the hypothesis of a common cell lineage for neurons and bipotential 0-2A progenitor cells and suggest that neuromodulin plays a more general role in plasticity during development of the central nervous system. The expression of neuromodulin in secondary cultures of newborn rat oligodendrocytes and its absence in type 1 astrocytes was confirmed by Northern blot analysis of isolated total RNA from these different types of cells using a cDNA probe for the neuromodulin mRNA and by Western blot analysis of the cell extracts using polyclonal antibodies against neuromodulin. The properties of the neuromodulin protein in cultured oligodendrocytes and neuronal cells have been compared. Although neuromodulin in oligodendrocytes is soluble in 2.5% perchloric acid like the neuronal counterpart it migrates essentially as a single protein spot on two-dimensional gel electrophoresis whereas the neuronal antigen can be resolved into at least three distinct protein spots. To obtain precise alignments of the different neuromodulin spots from these two cell types, oligodendrocyte and neuronal cell extracts were mixed together and run on the same two-dimensional gel electrophoresis system. Oligodendroglial neuromodulin migrates with a pI identical to the basic forms of the neuronal protein in isoelectric focusing gel. However, the glial neuromodulin shows a slightly lower mobility in the second dimensional lithium dodecyl sulfate-PAGE than its neuronal counterpart. As measured by 32Pi incorporation, neuromodulin phosphorylation in oligodendrocytes is dramatically increased after short-term phorbol ester treatments, which activate protein kinase C, and is totally inhibited by long-term phorbol ester treatments, which downregulates protein kinase C, thus confirming its probable specific in vivo phosphorylation by protein kinase C. In primary cultures of neuronal cells, two of the three neuromodulin spots were observed to be phosphorylated with an apparent preferential phosphorylation of the more acid forms.

Animals↗
PubMed1990

Neurogranin: immunocytochemical localization of a brain-specific protein kinase C substrate.

The developmental expression and the cellular localization of neurogranin (formerly designated p17), a brain-specific protein kinase C (PKC) substrate, were investigated. The developmental expression of neurogranin was studied by immunoblotting of rat brain and neuronal cell-culture extracts using neurogranin polyclonal antibodies. Neurogranin synthesis was found to be developmentally regulated, with no expression in the embryonic and neonatal period and an abrupt increase between 2 and 3 weeks of age. By immunohistochemistry, neurogranin was found essentially in the adult rat telencephalon, specifically located in the cell bodies and dendritic processes of neurons of the cerebral cortex, hippocampus, striatum, and a few other discreet areas. Neurogranin immunoreactivity was nearly absent in the thalamus, cerebellum, and brain stem. The late developmental expression and the dendritic localization of neurogranin in neurons are 2 features that also characterize the type I PKC isozyme. The specific localization of the protein in integrative areas of the rat brain suggests a highly specialized function of neurogranin in the CNS. A possible role for neurogranin in the transduction of the PKC activation signals at the postsynaptic level is suggested.

Animals↗
PubMed1989

The Ca2+-binding sequence in bovine brain S100b protein beta-subunit. A spectroscopic study.

Conformational changes in the beta-subunit of the bovine brain Ca2+-binding protein S100b (S100-beta) accompanying Ca2+ binding were investigated by analysis of the spectroscopic properties of the single tyrosine residue (Tyr17 beta) and flow-dialysis binding experiments. S100-beta binds Ca2+ sequentially at two sites to change the conformation of the protein. The first Ca2+ ion binds to site II beta, a typical Ca2+-binding site in the C-terminal region, and it does not significantly perturb the proximal environment of Tyr17 beta. After the first site is occupied, another Ca2+ ion binds to the N-terminal Ca2+-binding site, I beta, and strengthens a hydrogen bond between Tyr17 beta and a neighbouring carboxylate acceptor group, which results in a large increase in the Tyr17 beta fluorescence spectrum half-width and a positive absorption and c.d. signal between 290 and 275 nm. Ca2+ binding to the S100b.Zn2+6 complex, studied by flow-dialysis and fluorescence measurements showed that, although Zn2+ ions increase the affinity of S100b protein for Ca2+, the Ca2+-binding sequence was not changed. Tb3+ (terbium ion) binding studies on the S100b.Zn2+6 complex proved that Tb3+ antagonizes only Ca2+ binding site II beta and confirmed the sequential occupation of Ca2+-binding sites on the S100b.Zn2+6 complex.

Animals↗
PubMed1989

Protein kinase C substrates from bovine brain. Purification and characterization of neuromodulin, a neuron-specific calmodulin-binding protein.

Although such solubility is uncommon among proteins generally, several bovine brain proteins were found to be soluble in 2.5% perchloric acid, and many of them were in vitro substrates for protein kinase C (Ca2+/phospholipid-dependent enzyme). Two of the perchloric acid-soluble brain proteins were purified, p43 and p17. P43 and p17 could be phosphorylated by protein kinase C only in the presence of Ca2+ and phospholipids and neither was a substrate for protein kinase II. P43 was subsequently identified as the neurospecific, calmodulin-binding protein, neuromodulin (also designated P-57, GAP43, B50, or F1) (Alexander, K. H., Wakim, B. T., Doyle, G. S., Walsh, K. A., and Storm, D. R. (1988) J. Biol. Chem. 263, 7544-7549). A rapid purification method for neuromodulin was developed taking advantage of its newly discovered property, solubility in 2.5% perchloric acid, and of its previously recognized calmodulin-binding property. Evidence was obtained that neuromodulin isolated from cytosolic extract exists as a mixture of molecular forms and that the Ca2+-binding S100 protein-beta discriminates among the different neuromodulin isoforms in forming covalent complexes via disulfide bridges; this discrimination may be explained by analogous differences observed between the NH2-terminal amino acid sequences of p57 and F1. Solubility in 2.5% perchloric acid was demonstrated for another rat brain protein kinase C substrate, p87. We suggest that perchloric acid solubility might be a common property of protein kinase C substrates.

Animals↗
PubMed1989

Modulation of ATPase activities in the central nervous system by the S-100 proteins.

The isomeric forms of bovine S-100a and S-100b have been shown to stimulate ATPase activities in fractions enriched in myelin and mitochondria isolated from the Gerbil brain and for S-100b more effectively than for calmodulin in erythrocytes or skeletal muscle. In the presence of Ca2+, S-100a produced a slight increase of ATPase activity in the mitochondrial fraction. However, S-100b in the presence of Zn2+ almost doubled the ATPase activity in brain myelin. S-100a, or S-100b, with or without Ca2+ and Zn2+ respectively, had no effect on the ATPase activity in mitochondria of the Gerbil liver. The observations may indicate a "second messenger" role for S-100b in the presence of Zn2+ in the Schwann cell.

Adenosine Triphosphatases↗
PubMed1988

Interactions between the microtubule-associated tau proteins and S100b regulate tau phosphorylation by the Ca2+/calmodulin-dependent protein kinase II.

Binding between the microtubule-associated tau protein and S100b protein was demonstrated by affinity chromatography and cross-linking experiments and was manifested in the effect of S100b on tau protein phosphorylation by protein kinase II. All three expressions of the binding showed that S100b discriminates among the four species of tau, revealing for the first time that the different kinds of tau may differ functionally. Noncovalent interaction between tau and S100b depended on the presence of Ca2+ or Zn2+ and resulted in total inhibition of tau phosphorylation by protein kinase II. In the absence of reducing agent, covalent binding studies between Cys84 beta in the carboxyl-terminal region of the S100b-beta subunit and tau proteins confirmed interactions between the two proteins. It is suggested that the homologous calcium-binding domain that characterizes the carboxyl terminus of S100 and the tubulin subunit may be responsible for the common interaction of both proteins with tau proteins. The physicochemical relationship between S100 subunits and p11, the subunit of a substrate for tyrosine kinase, and their similarity in interaction with protein kinase substrates are discussed.

Animals↗
PubMed1988

Reinvestigation of the sulfhydryl reactivity in bovine brain S100b (beta beta) protein and the microtubule-associated tau proteins. Ca2+ stimulates disulfide cross-linking between the S100b beta-subunit and the microtubule-associated tau(2) protein.

Zn2+ and Ca2+ affect the conformation of bovine brain S100b (beta beta) protein and the exposure of its Cys-84 beta. Zn2+ binding to high-affinity sites of native S100b protected the sulfhydryl groups against the thiol-specific reagent 5,5'-dithiobis(2-nitrobenzoate) and antagonized the Ca2+-stimulated reactivity of Cys-84 beta toward the reagent. Spectroscopic studies on the fluorescence properties of labeled S100b with the fluorescent probes bimane and acrylodan at Cys-84 beta confirmed the antagonistic effect of Ca2+ and Zn2+ with respect to the conformational properties of the protein. Measurements of fluorescence dynamics on bimane-labeled S100b indicated that the slow monomer-dimer equilibrium that characterizes the apoprotein at micromolar concentrations was shifted to the monomer form in the presence of Zn2+, a fact that could explain the previously reported Zn2+-dependent increase of S100b protein affinity for calcium. The difference in the effects of Ca2+ and Zn2+ on the reactivity of Cys-84 beta in S100b was confirmed when we observed that Ca2+ and Zn2+ have opposite actions on the formation of disulfide bridges between Cys-84 beta of the S100b beta-subunit and sulfhydryl groups on the microtubule-associated tau(2) protein. Ca2+ stimulated the covalent complex formation whereas Zn2+ inhibited it. We suggest that Zn2+ may have a modulatory function on Cys-84 beta reactivity in the S100b beta-subunit in vivo. Two types of divalent complexes between tau(2) and beta-subunit were formed in the presence of Ca2+, an equimolar complex tau(2)-beta 1 and a complex of one molecule of tau(2) with two beta-subunits, tau(2)-beta 2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Phosphorylation of tau proteins to a state like that in Alzheimer's brain is catalyzed by a calcium/calmodulin-dependent kinase and modulated by phospholipids.

Calcium/calmodulin (CaM)-dependent protein kinases isolated from bovine and rat brains phosphorylate the microtubule-associated tau protein in the mode that shifts the mobility of tau in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (mode I). This mode of tau phosphorylation is the one that occurs abnormally in Alzheimer's lesions. Purified tau protein in solution can be phosphorylated by the Ca2+/CaM kinases maximally to about 50% of the total tau protein. Incorporation of one phosphate group per mol of tau is sufficient to shift the protein to a slower migrating electrophoretic band. Additional phosphate incorporation into the shifted tau proteins can occur depending on protein kinase concentration. In the presence of phosphatidylserine, tau proteins were phosphorylated to an extent of 100% at a tau: phosphatidylserine ratio of 20. Phosphatidylethanolamine also stimulated tau phosphorylation by Ca2+/CaM kinase and phosphatidylinositol was found to be a potent inhibitor of tau protein phosphorylation. The direct observation that tau proteins interact with phospholipids such as phosphatidylethanolamine and phosphatidylinositol, resulting in a smearing of the protein band on sodium dodecyl sulfate-gel electrophoresis, supports the possibility that tau protein may interact with phospholipid membranes in vivo and that tau protein phosphorylation could be modulated by the phospholipid composition of the membranes with which tau interacts.

Alzheimer Disease↗
PubMed1987

Separation of the different microtubule-associated tau protein species from bovine brain and their mode II phosphorylation by Ca2+/phospholipid-dependent protein kinase C.

Bovine brain tau protein (tau) consists of four closely related phosphoproteins named tau 1, tau 2, tau 3 and tau 4, that range in size from 55 to 68 kDa (as determined by gel electrophoresis). Here we report an improved large-scale purification method for tau protein and the separation of the four individual tau protein species. The separation of the individual tau protein was accomplished by two chromatographic techniques: hydroxyapatite chromatography allowed the separation of two pairs of tau protein (tau 1 and tau 3) and (tau 2 and tau 4); fast protein liquid chromatography on a Mono Q column at basic pH achieved the resolution of the individual tau protein species in each pair derived from hydroxyapatite columns. Chromatography on the Mono Q column revealed that tau protein possesses previously unrecognized, highly reactive sulfhydryl groups that may oxidize to form intermolecular disulfide bridges. The isolation of individual species of tau in substantial quantities permitted an improved amino acid analysis that demonstrated the occurrence of cysteine and tryptophan in the protein. The availability of individual tau protein species greatly simplified the analysis for mode II phosphorylation of tau, which was found to be catalyzed by the calcium/phospholipid-dependent protein kinase C. The mode II phosphorylation of tau by protein kinase C was not associated with a mobility shift for tau protein in SDS-polyacrylamide gel electrophoresis, in contrast to mode I phosphorylation of tau by the Ca2+/calmodulin-dependent kinase, which produces a substantial shift in mobility.

Amino Acids↗
PubMed1987

Comparison of S100b protein with calmodulin: interactions with melittin and microtubule-associated tau proteins and inhibition of phosphorylation of tau proteins by protein kinase C.

To gauge similarities between S100b protein and calmodulin, interactions were observed between S100b and melittin and between S100b and tau, the microtubule-associated proteins. The interaction of melittin with S100b protein in the presence and absence of calcium was studied by fluorescence polarization, UV difference spectroscopy, and sulfhydryl derivatization. Whether calcium was present or not in the solution, melittin and S100b form a complex of molar ratios up to 2:1. Further binding of melittin occurred, but it resulted in precipitation of S100b, as is true of the corresponding case of melittin binding to calmodulin. In the absence of calcium, the interaction of melittin and S100b shielded the tryptophan (Trp) of the former protein and exposed cysteine-84 beta (Cys-84 beta) of the latter protein, leaving the tyrosine-16 beta (Tyr-16 beta) of S100b unaffected. Calcium addition to the complex partially restored the exposure of Trp of melittin and caused changes in the environment of Tyr-16 beta (unlike the environmental changes induced for Tyr-16 beta by calcium in the absence of melittin). The conformational changes induced in S100b by interaction with melittin increased its affinity for calcium and offset the inhibition of calcium binding otherwise observed in the presence of potassium ions. This corroborated the previous finding that S100b affinity for calcium greatly depends on the protein conformation. The phenomena described above are similar to the interactions of melittin with calmodulin and thus suggest that S100b and calmodulin have a common structural domain not only that binds melittin but also that may interact with common target proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗