Purification and characterization of the catalytic subunit of cAMP-dependent protein kinase from the bivalve mollusc Mytilus galloprovincialis

Cao, Jesús; Fernández, Montserrat; Vázquez-Illanes, M D; Ramos-Martı́nez, Juan Ignacio; Villamarín, J. Antonio

doi:10.1016/0305-0491(95)00016-2

Cited by 16 publications

(11 citation statements)

References 51 publications

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“…Work from 1972 to 1988 showed that agents that relaxed catch (e.g., serotonin) increased muscle cAMP levels, and that chemical treatments that increased cAMP relaxed catch {Achazi, 1979 36504 /id; Köhler, 1980 36506 / id;Achazi, 1974 35209 /id;Cole, 1972 449 /id;Gies, 1986 36300 /id;Gies, 1988 23435 / id;Marchand-Dumont, 1975 36296 /id;Matsuura, 1984 480 /id;Painter, 1982 54 /id;Pfitzer, 1982 23455 /id}. Subsequent work isolated and characterized the regulatory and catalytic subunits of the cAMP-dependent protein kinase (Cao et al, 1995a(Cao et al, ,b, 1996Rodríguez et al, 1998;Díaz-Enrich et al, 2003;Bardales et al, 2004;Béjar and Villamarín, 2006). The importance of protein phosphorylation was further emphasized by the observation that a calcineuron type phosphatase was required for catch (Castellani and Cohen, 1992).…”

Section: Unique Properties Due To Acto-myosin Interaction 1: Catchmentioning

confidence: 99%

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Hooper

Hobbs

Thuma

2008

Progress in Neurobiology

131

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This is the second in a series of canonical reviews on invertebrate muscle. We cover here thin and thick filament structure, the molecular basis of force generation and its regulation, and two special properties of some invertebrate muscle, catch and asynchronous muscle. Invertebrate thin filaments resemble vertebrate thin filaments, although helix structure and tropomyosin arrangement show small differences. Invertebrate thick filaments, alternatively, are very different from vertebrate striated thick filaments and show great variation within invertebrates. Part of this diversity stems from variation in paramyosin content, which is greatly increased in very large diameter invertebrate thick filaments. Other of it arises from relatively small changes in filament backbone structure, which results in filaments with grossly similar myosin head placements (rotating crowns of heads every 14.5 nm) but large changes in detail (distances between heads in azimuthal registration varying from three to thousands of crowns). The lever arm basis of force generation is common to both vetebrates and invertebrates, and in some invertebrates this process is understood on the near atomic level. Invertebrate actomyosin is both thin (tropomyosin:troponin) and thick (primarily via direct Ca ++ binding to myosin) filament regulated, and most invertebrate muscles are dually regulated. These mechanisms are well understood on the molecular level, but the behavioral utility of dual regulation is less so. The phosphorylation state of the thick filament associated giant protein, twitchin, has been recently shown to be the molecular basis of catch. The molecular basis of the stretch activation underlying asynchronous muscle activity, however, remains unresolved.

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Section: Unique Properties Due To Acto-myosin Interaction 1: Catchmentioning

confidence: 99%

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Hooper

Hobbs

Thuma

2008

Progress in Neurobiology

131

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“…The standard reaction mixture contained, in a total volume of 60 pl, 20 mM potassium phosphate pH 6.8, 1.5 mM MgCl,, 5.0 mg/ml histone (type IIA, Sigma), 1 pM cAMP (when used), 0.37 mM [y-"PIATP (125-150 cpndpmol) and the sample containing protein kinase activity. After 5 min at 30"C, the reaction was stopped by adding 1 ml ice-cold 10% trichloroacetic acid and the mixture processed as described previously [9].…”

Section: Methodsmentioning

confidence: 99%

“…All other chemicals were of the highest purity available from commercial sources. Catalytic (C) subunit of CAMP-dependent protein kinase from mussel was purified as described in [9] ; catalytic and regulatory (RII) subunits from porcine heart were purified by the methods of Reimann and Beham [lOJ and Rannels et al [l 11 respectively.…”

Section: Methodsmentioning

confidence: 99%

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Characterization of a cAMP-Binding Protein from the Bivalve Mollusc Mytilus galloprovincialis

Cao¹,

Ramos-Martı́nez²,

Villamarı́n³

1995

Eur J Biochem

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Three CAMP-binding proteins have been identified by photoaffinity labeling with S-azido["?P]cAMP and purified from the mantle tissue of the sea mussel Mytilus ,eullo~,rovinc.ialis. Their molecular masses, determined by SDSPAGE, were 54, 42 and 37 kDa. The purified 54-kDa protein, which had two CAMPbinding sitedmonomer, was judged to be a regulatory (R) subunit of CAMP-dependent protein kinase since it re-associated with and inhibited purified catalytic (C) subunit of this enzyme from mussel, in the absence but not in the presence of CAMP. The molecular mass of the complex between Mytilus CAMPbinding protein and C subunit, estimated by analytical gel-filtration, was 220 kDa, a value which agrees with a RZCZ stoichiometry for the mussel CAMP-dependent protein kinase holoenzyme. On the basis of the elution pattern from DEAE-cellulose chromatography and its ability to be phosphorylated by purified C subunit of CAMP-dependent protein kinase, the 54-kDa protein could be classified as a type 11 regulatory subunit. Furthermore, no mobility shift on SDSlPAGE upon phosphorylation/dephosphoryIation of Myrilus protein was observed, a similar behaviour to that shown by the mammalian RIIP isoform. The 42-kDa and 37-kDa proteins, which were recognized by a specific antiserum against the 54-kDa protein and fail to be phosphorylated by M.ytiZu.s C subunit, are probably products generated by proteolysis of the 54-kDa protein, although they were shown even when inhibitors of the major types of proteases were used.Keyvords: CAMP-binding protein; CAMP-dependent protein kinase; regulatory subunit; protein phosphorylation ; Mytilus.For intertidal marine invertebrates, such as some bivalve molluscs, access to environmental oxygen is intermitent, so that these organisms are subjected to cyclic hypoxia periods. It is well described that, during hypoxia, a metabolic rate dcprcssion takes place, especially of glycolytic pathway [I]. One of the mechanisms involved in this biochemical adaptation seems to be the covalent modification (phosphorylation/dephosphorylation) of the key regulatory glycolytic enzymes : 6-phosphofructo-Ikinase, 6-phusphofructo-2-kinase and pyruvate kinase 12-41, However, despite the fact that certain neuroendocrine factors affecting the gycogen metabolism have been recently recognized in the sea mussel Mytilus edu1i.s [5, 61, their structure remains unknown and, so far, very little information is available about the mechanisms of intracellular signalling in these organisms.In our laboratory, we have proved that, in the sea mussel Mytilus ~~mllo~~r~~viiir~ialis, two key glycolytic enzymes, 6-phosphofructo-1 -kinase and 6-phosphofructo-2-kinase, are activated when phosphorylated in vitro by CAMP-dependent protein kinase [7, XI. On the other hand, it has also been reported that incubation of a dialyzed homogenate from mussels with cAMP C o t . r~. s~~o n d~~r i c~~ r o J. A. Villamarin,

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“…In this sense, data suggesting an in-volvement of the system mediated by adenosine 3Ј,5Ј-cyclic monophosphate (cAMP) are available. The catalytic subunit of cAMP-dependent protein kinase (EC 2.7.1.37) was isolated from mantle tissue of M. galloprovincialis (10). This enzyme was proved to catalyze the PFK-l phosphorylation at the level of a serine residue (8).…”

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confidence: 99%

Measurement of Adenosine 3′,5′-Cyclic Monophosphate and Guanosine 3′,5′-Cyclic Monophosphate in Mussel (Mytilus galloprovincialis Lmk.) by High-Performance Liquid Chromatography with Diode Array Detection

Enrich

Villamarı́n

Martinez

et al. 2000

Analytical Biochemistry

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Purification and characterization of the catalytic subunit of cAMP-dependent protein kinase from the bivalve mollusc Mytilus galloprovincialis

Cited by 16 publications

References 51 publications

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Invertebrate muscles: Thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

Characterization of a cAMP-Binding Protein from the Bivalve Mollusc Mytilus galloprovincialis

Measurement of Adenosine 3′,5′-Cyclic Monophosphate and Guanosine 3′,5′-Cyclic Monophosphate in Mussel (Mytilus galloprovincialis Lmk.) by High-Performance Liquid Chromatography with Diode Array Detection

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