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P J Coggins

Publications and source records attributed to P J Coggins.

13 recordsLinked to original sources
PubMed1994

Detergents and peptides alter proteolysis and calmodulin binding of B-50/GAP-43 in vitro.

The neuronal growth-associated protein B-50/GAP-43 is a substrate for protein kinase C, binds to calmodulin in a calcium-independent manner, and in vitro is subject to an endogenous and chymotrypsin-mediated hydrolysis in the vicinity of the single kinase C phosphorylation site. All of these processes can be influenced by corticotrophin (ACTH). In the present study we have investigated whether these biochemical interactions involving B-50 could have common structural determinants. Chymotryptic digestion of B-50 in the presence or absence of a nonionic detergent and ACTH demonstrated that hydrolysis is potentiated by a lipid-like environment that primarily affects the protein rather than the protease or the peptide. Furthermore, this lipid dependency appears to extend to the binding of dephosphorylated B-50 to calmodulin, which appears to occur only in the presence of a nonionic detergent or lipid and the absence of calcium. A structure-activity study for ACTH-mediated inhibition of B-50 proteolysis by an endogenous protease that copurifies with B-50 in a detergent extract of synaptosomal plasma membranes showed that ACTH1-24, ACTH5-24, ACTH5-16, dynorphin, and corticostatin inhibited the conversion of rat B-50 to B-5041-226. In contrast, ACTH7-16, Org2766, and neurotensin had no detectable effect on B-50 proteolysis at concentrations of 10 and 50 microM.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1993

Neurogranin, a B-50/GAP-43-immunoreactive C-kinase substrate (BICKS), is ADP-ribosylated.

Neurogranin is a neurone-specific, B-50-immunoreactive C-kinase substrate that has limited homology to, but considerable biochemical similarity to B-50/GAP43. The most significant differences between these two proteins are their cellular localisation and molecular mass (Neurogranin, 7.5 kDA cytosolic; and B-50, 25 kDa membranal). An understanding of the similarities and differences between Neurogranin and B-50 may facilitate the elucidation of their hitherto elusive functions in the nervous system. The results of the present study demonstrate that, in common with B-50, Neurogranin is a substrate for ADP-ribosyltransferase. This finding is discussed with regard to the concept of molecular flexibility of B-50-like proteins as the basis of their putatively diverse roles in the nervous system.

Adenosine Diphosphate Ribose↗
PubMed1993

ADP-ribosylation of the neuronal phosphoprotein B-50/GAP-43.

The neuronal phosphoprotein B-50/GAP-43 is associated with growth and regeneration within the nervous system and its posttranslational status can be correlated with its cellular localization during growth and regeneration. Recently, B-50 has been shown to interact with certain G protein subunits. Regulation of G protein-mediated signal transduction may involve ADP-ribosylation in vivo. In the present study we have demonstrated that B-50 is a substrate for endogenous ADP-ribosyltransferases. The results are discussed with respect to the possible interaction of B-50 with G proteins, but also with regard to the posttranslational modification of B-50 by all major regulatory mechanisms that act at, or through, the neuronal membrane.

Adenosine Diphosphate Ribose↗
PubMed1991

B-50 (GAP-43): biochemistry and functional neurochemistry of a neuron-specific phosphoprotein.

The biochemistry and functional neurochemistry of the synaptosomal plasma membrane phosphoprotein B-50 (GAP-43) are reviewed. The protein is putatively involved in seemingly diverse functions within the nervous system, including neuronal development and regeneration, synaptic plasticity, and formation of memory and other higher cognitive behaviors. There is a considerable amount of information concerning the spatial and temporal localization of B-50 (GAP-43) in adult, fetal, and regenerating nervous tissue but far less is known about the physical chemistry and biochemistry of the protein. Still less information is available about posttranslational modifications of B-50 (GAP-43) that may be the basis of neurochemical mechanisms that could subsequently permit a variety of physiological functions. Hence, consideration is given to several plausible roles for B-50 (GAP-43) in vivo, which are discussed in the context of the cellular localization of the protein, significant posttranslational enzymes, and regulatory proteins, including protein kinases, phosphoinositides, calmodulin, and proteases.

Adrenocorticotropic Hormone↗
PubMed1990

Distribution of growth-associated protein, B-50 (GAP-43) in the mammalian enteric nervous system.

The presence of the growth-associated protein, B-50 (also known as GAP-43) was investigated in the adult mammalian enteric nervous system. The small intestine of rat, ferret and human was examined by immunohistochemistry. Dense B-50-like immunoreactivity was localized in nerves throughout the wall of the rat, ferret and human small intestine, notably in the myenteric and submucous plexuses, where in the ferret ileum it co-localized with vasoactive intestinal polypeptide-immunoreactive fibre groups. Material with the biochemical and immunological characteristics of rat B-50 was extracted from the rat ileum. In-situ hybridization demonstrated that enteric neurons express B-50. These findings are consistent with a role for B-50 in the documented plasticity of the adult enteric nervous system.

Animals↗
PubMed1990

Binding of the neuronal protein B-50, but not the metabolite B-60, to calmodulin.

Protein kinase C phosphorylates the neurone-specific protein B-50 at a single Ser41 residue, which is also the point for a major proteolytic cleavage in vitro, and probably in vivo, that produces a B-50 phosphorylation-inhibiting N-terminal fragment and a large C-terminal metabolite B-60 (B-50(41-226]. The intact purified protein will bind to calmodulin in the absence of calcium, but the interaction has an absolute requirement for dephospho-B-50. In an attempt to unify two aspects of B-50 biochemistry, we have examined the interaction of B-50 binding to calmodulin and B-50 proteolysis. HPLC- and affinity-purified B-50 bound to calmodulin, but purified B-60 did not. To ensure that this effect was not due to the phosphorylation state of pure, isolated B-60, the metabolite was generated in vitro using a Triton extract of synaptosomal plasma membranes, which contains the as yet uncharacterized B-50 protease. B-60 derived from dephospho-B-50 also failed to bind calmodulin. The results demonstrate a direct connection between B-50 binding to calmodulin and B-50 proteolysis. The position of the proposed calmodulin-binding domain within intact B-50 is discussed in light of the failure of calmodulin to bind B-60.

Animals↗
PubMed1989

Evidence for a single protein kinase C-mediated phosphorylation site in rat brain protein B-50.

The neuronal protein B-50 may be involved in diverse functions including neural development, axonal regeneration, neural plasticity, and synaptic transmission. The rat B-50 sequence contains 226 amino acids which include 14 Ser and 14 Thr residues, all putative sites for phosphorylation by calcium/phospholipid-dependent protein kinase C (PKC). Phosphorylation of the protein appears to be a major factor in its biochemical and possibly its physiological activity. Therefore, we investigated rat B-50 phosphorylation and identified a single phosphorylated site at Ser41. Phosphoamino acid analysis eliminated the 14 Thr residues because only [32P]Ser was detected in an acid hydrolysate of [32P]B-50. Staphylococcus aureus protease peptide mapping produced a variety of radiolabelled [32P]B-50 products, none of which had the same molecular weights or HPLC retention times as several previously characterized fragments. Indirect confirmation of the results was provided by differential phosphorylation of major and minor forms of B-60 that have their N-termini at, or C-terminal to, the Ser41 residue and are the major products of specific B-50 proteolysis. Only those forms of B-60 that contained the Ser41 residue incorporated phosphate label. The results are discussed with reference to the substrate requirements for B-50 phosphorylation by PKC and the proposed structure of the B-50 calmodulin binding domain.

Amino Acid Sequence↗
PubMed1987

High affinity specific binding of the thyrotrophin releasing hormone metabolite histidylproline to rat brain membranes.

Histidylproline a metabolite of Thyrotrophin Releasing Hormone specifically binds to both high and low affinity sites in fresh rat brain membranes. Characterisation of the high affinity site under optimal conditions demonstrated an equilibrium dissociation constant (KD) of approximately 9nM and maximum binding capacity of approximately 120 fmols/mg protein. Kinetic analysis of [3H]-His-Pro binding is limited by low binding density, instability of the high affinity site and rapid degradation of the radioligand. The thiol blocking reagent pHydroxymercuriphenylsulphonic acid (HMPS) inhibited [3H]-His-Pro degradation but also reduced binding of the peptide to membranes. The results are discussed with reference to the lack of specific binding sites in brain for the proposed neuropeptide and TRH metabolite cyclo(His-Pro).

Animals↗
PubMed1986

Histidyl-proline, a rapidly degraded metabolite of thyrotropin releasing hormone, has behavioural activity.

The effects of intracerebroventricular (ICV) administration of histidyl-proline (His-Pro; 0, 1, 10 and 100 micrograms/rat), a rapidly degraded metabolite of thyrotropin releasing hormone (TRH), were assessed using three behavioural paradigms. Locomotor activity was significantly increased over 30, but not 90, min following administration of the highest dose. This stimulatory effect was more apparent when naive rats were tested in an open field without any prior exposure to the apparatus; the effect was attenuated on a second exposure. His-Pro treated subjects also tended to prefer the area near the walls of the apparatus. The effects of the dipeptide on visual perception were studied, using a two-choice task. A low but not high (10, versus 100 micrograms/rat) dose of His-Pro increased response switching, but did not affect response repetition, latencies or percent correct performance. The results indicate that His-Pro can alter behaviour by affecting motor output, as well as emotional processes.

Animals↗
PubMedDate not supplied

Thyrotrophin releasing hormone degradation by rat synaptosomal peptidases: production of the metabolite His-Pro.

The dipeptide, His-Pro, is the major product of the degradation of TRH by rat synaptic membranes in vitro. A small amount of His-Pro is also formed from TRH by the synaptosomal soluble fraction. From inhibitor studies, the main route to His-Pro appears to involve removal of the pGlu residue by membrane-bound metal-dependent pyroglutamylaminopeptidase followed by deamidation. The deamidation step is not mediated by proline endopeptidase (EC3.4.21.26) nor dipeptidylpeptidase-IV (EC3.4.14.5) since it is insensitive to bacitracin and diprotin-A, and may therefore involve a novel membrane-bound TRH metabolizing enzyme. His-Pro is degraded rapidly by the soluble synaptosomal fraction, presumably by prolidase (EC3.4.13.9) and more slowly by the synaptic membrane fraction.

Animals↗
PubMedDate not supplied

Corticotropin (ACTH) inhibits the specific proteolysis of the neuronal phosphoprotein B-50/GAP-43.

The modulatory action of corticotropin (ACTH) on the specific proteolysis of B-50 to B-60 [B-50(41-226)] was investigated using a previously characterized synaptosomal membrane extract [ASP(57-82)] that was highly enriched in B-50, B-50 kinase activity and a B-50 protease. In common with the previously described inhibitory action on B-50 phosphorylation at Ser41, ACTH(1-24) inhibited B-50 proteolysis in a concentration and structurally dependent manner, while ACTH(1-10) and ACTH(11-24) or a combination of both peptides were ineffective. Structural similarities between segments of ACTH(1-24) and B-50(40-55) may explain the inhibitory action of the peptide on B-50 phosphorylation and proteolysis. In addition, calmodulin, which binds to the N-terminus of the dephosphorylated form of the protein, also protected B-50 from degradation.

Adrenocorticotropic Hormone↗