The structural basis and biological role of protein-protein interactions: a multidisciplinary approach  Page description

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Details of project

  
Identifier
49812
Type NI
Principal investigator Gráf, László
Title in Hungarian A fehérje-fehérje kölcsönhatások szerkezeti alapjai és biológiai szerepük: multidiszciplináris megközelítés
Title in English The structural basis and biological role of protein-protein interactions: a multidisciplinary approach
Panel Genetics, Genomics, Bioinformatics and Systems Biology
Department or equivalent Department of Biochemistry (Eötvös Loránd University)
Participants Gombos, Linda
Hegyi, György
Hetényi, Csaba
Hódi, Zsuzsanna
Kardos, József
Kellermayer, Miklós Sándor Zoltán
Láng, András
Málnási Csizmadia, András
Németh, Attila
Nyitray, László
Pál, Gábor
Patthy, András
Perczel, András
Szilágyi, László
Venekei, István
Starting date 2005-01-01
Closing date 2008-12-31
Funding (in million HUF) 87.000
FTE (full time equivalent) 17.10
state closed project





 

Final report

  
Results in Hungarian
Az ELTE Biokémiai Tanszék tudományos kutatásainak tengelyében évtizedek óta a fehérjék szerkezetének, funkciójának és a fehérjék közötti kölcsönhatások szerkezeti hátterének és biológiai jelentőségének felderítése áll. Valamennyi, az elmúlt négyéves pályázati ciklusban vállalt feladatunk teljesítése a fenti célokat szolgálta. A vezető kutató megítélése szerint a pályázat támogatásával elért legkiemelkedőbb eredményeink, a vállalt témák sorrendjében a következők voltak: 1) Megállapítottuk, hogy a primata-specifikus tripszin 4 egyik, feltehetően biológiai szubsztrátja a mielin bázikus fehérje és modellt dolgoztunk ki a humán tripszinogén 4 asztroglia sejteken belüli transzportjának és aktivációjának követésére 2) Hazánkban elsőként állítottuk be a fágbemutatás módszerét, melynek segítségével az eredetitől eltérő specifitású proteáz inhibitorokat állítottunk elő. 3) Megállapítottuk, hogy a miozin II motorfehérje regulációs képességének az az előfeltétele, hogy aproximális kétláncú coiled-coil szerkezet instabil legyen. 4) Felderítettük a miozin II specifikus inhibitorának, a blebbistatinnak a működésmechanizmusát. 5) Kifejlesztettünk egy új tranziens kinetikai módszert, a „temperature-jump/stopped flow-t a módszer alkalmazásához szükséges berendezéssel együtt.
Results in English
For decades the focus of scientific interest of the Biochemistry Department, Eötvös Loránd University, Budapest has been the investigation of the structural basis and biological significance of protein-protein interactions. Our research efforts during the last 4 years were to achieve specific goals along this line. In the view of the principal investigator of this grant the most outstanding scientific results achieved, or discoveries made by using the financial means of the grant are as follows: 1) We provided indirect evidence that one of the potential pathological substrate of human trypsin 4 might be myelin basic protein. Furthermore, we worked out a model to follow the transport and activation of human trypsinogen 4 within human astroglia cells. 2) For the first time in Hungary we introduced the methodology of phage-display in our department and by using this method we succeeded in producing serine protease inhibitors with altered specificity. 3) Evidence was provided that the instability of the proximal two-chain coiled structure in myosin II plays an important regulatory role in the myosin functioning 4) The mechanism of action of the inhibitor of myosin II, blebbistatin was explored. 5) We developed and set up a new method and apparatus to perform „temperature-jump/stopped flow” transient kinetics experiments.
Full text https://www.otka-palyazat.hu/download.php?type=zarobeszamolo&projektid=49812
Decision
Yes





 

List of publications

  
Kardos, J., Okuno, D., Kawai, T., Shimizu, Y., Yumoto, N., Kitagawa, T., Závodszky, P., Naiki, H., and Goto, Y.: Structural studies reveal that the diverse morphology of ¦Â2-microglobulin aggregates .is a reflection of different molecular architectures., BBA 1753, 108-120., 2005
Farkas, L. Debreczeni, J. Harmat, V., Hetényi, Cs., Hajdú, I., Závodszky, P., Kohama, K., and Nyitray, L: Ca2+ inhibits the activity of a myosin II by binding to an EF-hand of a calmodulin-like light chain and altering the dynamics of the neck region., J. Muscle Res. Cell Motility. 26:74, 2005
Hódi, Zs., Németh, A., Kovács, E., Hetényi, Cs., Bodor, A., Perczel, A., and Nyitray, L.: The dynein light chain binds to a non-coiled-coil tail domain of myosin-Va that includes an alternatively spliced exon coding for three amino acid residues., FEBS J. 272 (suppl.1):335, 2005
Kondrák, M., Kutas, J., Szenthe, B., Patthy, A., Bánfalvy, Zs., Nádasy, M., Gráf, L., Asbóth, B.: Inhibition of Colorado potato beetle larvae by a locust proteinase inhibitor peptide expressed in potato., Biotechnology Letters 12, 829-834., 2005
Malnasi-Csizmadia, A., Dickens, J.L., Zeng, W., Bagshaw, C.R.: Switch movements and the myosin crossbridge stroke., J Muscle Res Cell Motil. 26(1), 31-37., 2005
Szenthe, B., Frost, C., Szilágyi, L., Patthy, A. Naudé, R., Gráf, L.: Cloning and expression of ostrich trypsinogen: an avian trypsin with a highly sensitive autolysis site., Biochim. Biophys. Acta 1748, 35-42., 2005
Fodor, K., Harmat, V., Kardos, J., Antal, J., Hetényi, C., Perczel, A., Szenthe, B., Gáspári, Z., Katona, G., Gráf, L.: Conformational adaptation of a canonical protease inhibitor upon its binding to the target protease increases specificity., FEBS J. 272, (s1)., 2005
Medveczky, P., Siklódi, E., Tóth, J., Patthy, A., Gallatz, K., Németh, P., Palkovits, M., Gráf, L., Szilágyi, L.: Trypsinogen 4 with a 28 amino acid leader peptide on its N- terminus is the predominant form of the enzyme in human brain., FEBS J. 272, (s1)., 2005
Medveczky, P., Antal, J., Patthy, A., Kékesi, K., Juhász, G., Szilágyi, L., Gráf, L.: Human trypsin 4 selectively cleaves myelin basic protein: Is this brain protease involved in the pathomechanism of multiple sclerosis?, FEBS J. 272, (s1)., 2005
Gombos, L., Tóth J., Medveczky P., Málnási Csizmadia A., Szilágyi L.: Comparative kinetic study on S2' trypsin variants., FEBS J. 272, (s1)., 2005
Tóth, J., Gombos, L., Siklódi, E., Németh, P., Palkovits, M., Szilágyi, L., Gráf, L.: Regional distribution of human trypsinogen 4 in human brain determined at mRNA and protein level., FEBS J. 272, (s1)., 2005
Gráf, L., Fodor, K., Szenthe, B., Perczel, A., Gáspári, Z., Harmat, V., Hetényi, C., Antal, J., Kardos, J., Katona, G., Patthy, A.: Protease inhibitors of the grasshopper family: structure, molecular flexibility, specificity and mechanism of action., FEBS J. 272, (s1)., 2005
Marokházi, J., Felföldi, B., Mihala, N., Hudecz, F., Patthy, A., Fodor, A., Gráf, L., Venekei, I.: Identification of cleavage site and natural substrate specificity of PrtA, a serralysin-type metalloprotease from the entomopathogenic microorganism Photorhabdus., FEBS J. 272, (s1)., 2005
Kawamichi, H., Hino, M., nakamura, A., Tanaka, H., Frakas, L., Nyitray, L., Kohama, K.: calcium inhibition of Physarium myosin as examined by the recombinant heavy mero-myosin, Adv. Exp. Med. Biol. 592:265-272., 2007
Süveges, D. Németh, A., Gáspári, Z., Tóth, G., and Nyitray, L.: Helical tail fragments of myosin VI exist as monomeric, highly flexible structures., J. Muscle Res. Cell Motility. 26:74, 2005
Fodor, K., Harmat, V., Neutze, R., Szilágyi, L., Gráf, L., Katona, G.: Overview of the acylation phase of serine protease catalysis at atomic resolution: The role of enzyme:substrate hydrogen bonds, 2006 Feb 21;45(7):2114-21., 2006
Hetenyi, C., Paragi, G., Maran, U., Timar, Z., Karelson, M., Penke, B.: Combination of a modified scoring function with two-dimesional descriptors for calculation of binding affinities of bulky, flexible ligands to proteins., J Am Chem Soc. 2006 Feb 1;128(4):1233-9., 2006
Medveczky, P., Antal, J., Patthy, A., Kékesi, K., Juhász, G., Szilágyi, L., Gráf, L.: Myelin basic protein, an autoantigen in multiple sclerosis, is selectively processed by human trypsin 4, FEBS Lett. 2006 Jan 23;580(2):545-52., 2006
Szenczi, A., Kardos, J., Medgyesi, G., Zavodszky, P.: The effect of solvent environment on the conformation and stability of human polyclonal IgG in solution., 2006 Mar;34(1):5-14., 2006
Tóth, J., Medveczky, P., Szilágyi, L., Gráf, L.: Central nervous system serine proteases in health and disease., Handbook of neurochemistry and molecular neurobiology, 3rd edition. (Lajtha, A., ed.) Springer-Verlag, 2007
Hódi, Zs. Németh, A., Hetényi, Cs., Bodor, A., Perczel, A. and Nyitray, L.: Alternatively spliced exon B of myosin Va is essential for binding of the tail light chain shared by dynein., J. Muscle Res. Cell Motility. 26:73, 2005
Zeng W, Seward HE, Malnasi-Csizmadia A, Wakelin S, Woolley RJ, Cheema GS, Basran J, Patel TR, Rowe AJ, Bagshaw CR.: Resonance energy transfer between green fluorescent protein variants: complexities revealed with myosin fusion proteins., Biochemistry 2006, 45(35):10482-91., 2006
Kintses B, Simon Z, Gyimesi M, Toth J, Jelinek B, Niedetzky C, Kovacs M, Malnasi-Csizmadia A.: Enzyme kinetics above denaturation temperature: a temperature-jump/stopped-flow apparatus., Biophys J. 2006;91(12):4605-10, 2006
Tóth J., Simon Z., Medveczky P., Gombos L., Jelinek B., Szilágyi L., Gráf L., Málnási-Csizmadia A.: Sitedirected mutagenesis at position 193 of Human trypsin 4 alters the rate of conformational change during activation, Proteins 67:1119-1127, 2007
Yang, Y., Gourinath, S., Kovács, M., Nyitray, L., Reutzel, R., Himmel, D.M., O'Neall-Hennessey, E., Reshetnikova, L., Szent-Györgyi, A.G., Brown, J.H., and Cohen, C.: Rigor-like structures of muscle myosins reveal key mechanical elements in the transduction pathways of this allosteric motor., Structure 15:553-564, 2007
Németh Attila, Süveges Dániel, Kovács Mihály, Nyitray László: Miozin II motorfehérjék regulációjának kinetikai vizsgálata., Biokémia 30(3):76, 2006
Süveges D., Tóth G., Hetényi Cs., Gáspári Z., Perczel A., Nyitray L: Egyszálú, stabil α-helikális elemek keresése és szerkezeti jellemzésük., Biokémia 30(3):78, 2006
Hódi Zs., Németh A., Kardos J., Radnai L., Hetényi Cs., Schlett K., Bodor A., Perczel A., Nyitray L.: Miozin Va: egy 3 aminosavat kódoló alternatív exon és egy dinein könnyű lánc., Biokémia 30(3):71, 2006
Beke T., Csizmadia G.I., Perczel A.: Theoretical Study on Tertiary Structural Elements of b-peptides: Nanotubes Formed from Parallel-Sheet-Derived Assemblies of b-Peptides, J. AM. CHEM. SOC. 2006, 128, 5158-5167, 2006
Pohl G., Beke T., Borbély J., Perczel A.: Prediction of Folding Preference of 10 kDa Silk-like Proteins Using a Lego Approach and ab Initio Calculations, J. AM. CHEM. SOC. 2006, 128,, 2006
Hudáky P., Perczel A.: A self-stabilized model of the Chymotrypsin Catalytic Pocket. The energy profile of the overall catalytic cycle, Proteins: Structure, Function and Bioinformatics, 2006, 62, 749-759, 2006
Hegyi G., Belágyi J.: Intermolecular cross-linking of F-actin alters the dynamics of its interaction with H-meromyosin in the weak-binding state., FEBS Journal 273 (2006) 1896-1905, 2006
Gaspari Z, Szenthe B, Patthy A, Westler WM, Graf L, Perczel A.: Local binding with globally distributed changes in a small protease inhibitor upon enzyme binding, FEBS J. 2006 Apr;273(8):1831-42., 2006
Németh, A., Medveczky, P., Tóth, J., Siklódi, E., Schlett, K., Patthy, A., Palkovits, M., Ovádi, J., Tőkési, N., Németh, P., Szilágyi, L., Gráf L.: Unconventional translation initiation of trypsinogen 4 at CUG codon with an N-terminal leucine: possible means to regulate gene expression., FEBS Journal 274: 1610-1620, 2007
Marokházi J., Mihala N., Hudecz F., Fodor A., Gráf L., Venekei I.: Cleavage site analysis of a serraysin-like protease, PrtA, from an insect pathogen Photorhabdus luminescens and development of a highly sensitive and specific substrate, FEBS Journal 274:1946-1956, 2007
Tóth, J., Siklódi, E., Medveczky, P., Gallatz, K., Németh, P., Szilágyi, L., Gráf, L., Palkovits, M.: Regional distribution of human trypsinogen 4 in human brain at mRNA and protein level, Neurochem. Res. 32:1423-1433, 2007
Gallatz, K., Medveczky, P.,Németh, P., Szilágyi, L., Gráf, L., Palkovits, M.: Human trypsin(ogen) 4-like immunoreactivity in the white matter of cerebral cortex and the spinal cord, Ideggyógyászati Szemle 60:18-23, 2007
Gallatz, K., Medveczky, P.,Németh, P., Szilágyi, L., Gráf, L., Palkovits, M.: Human trypsin(ogen) 4-like immunoreactivity in the white matter of cerebral cortex and the spinal cord, Ideggyógyászati Szemle 60:18-23, 2007
Jelinek, B., Katona, G., Fodor, K., Venekei, I., Gráf, L.: The crystal structure of a trypsin-like mutant chymotrypsin: the role of position 226in the activity and specificity of S189D chymotrypsin, Protein. J. 27:79-87, 2008
Hódi, Zs., Németh, A., Radnai, L., Hetényi, Cs., Schlett, K., Bodor, A., Perczel, A., Nyitray, L.: Alternatively spliced exon B of myosin Va is essential for binding of the tail light chain shared by dynein, Biochemistry 45:12582-12595., 2007
Málnási-Csizmadia, A., Tóth, J.. Pearson, D.S., Hetényi, Cs., Nyitray, L., Geeves, M.A., Bagshaw, C.R., C.R., Kovács, M.: Selective perturbation of the myosin recovery stroke by point mutations at the base of the lever arm affects ATP hydrolysis and phosphate release, J. Biol. Chem. 282: 17658-17664., 2007
Hódi, Zs., Rapali, P., Radnai, L., Molnár, T., Szenes, Á., Kardos, J., Buday, L., Stafford, W.F., Nyitray, L.: The LC8 family of dynein light chains. multifunctional chaperon-like proteins, FEBS J. 274:106., 2007
Süveges, D., Gáspári, Z., Tóth, G., Nyitray, L.: Prediction of highly charged single alpha-helical elements in proteins, FEBS J. 274: 250., 2007
Gyimesi, M., Tsaturyan, A., Kellermayer, M., Málnási-Csizmadia, A.: Kinetic characterization of the function of myosin loop 4 in the actin-myosin interaction, Biochemistry, in press, 2007
Hudáky, P., Perczel, A.: pKa optimized catalysis in serine proteases, an Ab Intitio study on the catalytic His, Intern. J. Quantum Chemistry 107: 2178-2183, 2007
Gombos, L., Kardos, J., Patthy, A., Medveczky, P., Szilágyi, L., Málnási-Csizmadia, A. Gráf, L.:: Probing Conformational Plasticity of the Activation Domain of Trypsin: the Role of Glycine Hinges, Biochemistry 47: 1675-1684, 2008
Süveges, D., Gáspári, Z., Tóth, G., Nyitray, L.: Changes single a-helix: a verasatile protein natural motif, Proteins 74:905-9016, 2009
Felföldi, G. Marokházi, J., Képíró, M., Venekei, I.: Identification of natural target proteins indicates functions to a serralysin-type metalloprotease, PrtA, in antiimmune mechanisms, Applied and Environmental Microbiology, in press, 2009
Ángyán, A.F., Perczel, A., Pongor, S., Gáspári, Z.: Fast protein fold estimation from NMR-derived distance restraints, Bioinformatics 24: 272-275, 2008
Fodor, K., Harmat, V., Hetényi, C., Kardos, J., Antal, J., Perczel, A., Katona, G., Gráf, L.: Extended intermolecular interactions in a serine protease-canonical inhibitor complex account for strong and highly specific inhibition., J. Mol. Biol. 350, 156-169., 2005
Toth J, Gombos L, Simon Z, Medveczky P, Szilagyi L, Graf L, Malnasi-Csizmadia A.: Thermodynamic analysis reveals structural rearrangement during the acylation step in human trypsin 4 on 4-methylumbelliferyl 4-guanidinobenzoate substrate analogue., J Biol Chem. 2006, 281(18):12596-602., 2006
Perczel, A., Hudáky, P., Pálfi, V.K.: Dead-end street of protein folding: thermodynamic rationale of amyloid fibril formation, J. Am. Chem. Soc. in press, 2007
Szenthe B., Patthy A., Gáspári Z., Kékesi A.K., Gráf L., Pál G.: The Surface Reports What Lies Beneath: Combinatorial Phage-display Mutagenesis reveals Complex Networks of Surface-core Interactions in the Pacifastin Protease Inhibitor, J. Mol. Biol. 370:63-79, 2007
Tóth, J., Varga, B., Kovács, M., Málnási-Csizmadia, A., Vértessy, B.G.: Kinetic mechanism of human dETPase, an essential nucleotide pyrophosphatase enzyme, J. Biol. Chem. 282: 33572-33582, 2007
Jákó, É., Ittzés, P., Szenes, Á., Kun, Á., Szathmáry, E., Pál, G.: In silico detection of tRNA sequence features characteristic to aminoacyl-tRNA synthetase class membership, Nucleic Acid Research 35:5593-5609, 2007
Kinses, B., Gyimesi, M., Pearson, D.S., Geeves, M.A., zeng, W., Bagshaw, C.R., Málnási-Csizmadia, A.: Reversible movement of switch I loop of myosin determines actin interaction., EMBO J. 26: 265-274., 2007
Kardos, J., Harmat, V., Pallo, A., Barabás, O., Szilágyi, K., Gráf, L., Szabo, G.N., Goto, Y., Závodszky, P., Gál, P.: Revisiting the mechanism of autoactivation of the complement protease C1r in the C1 complex: Structure of the active catalytic region of C1r, Mol. Immunol. 45: 1752-1760, 2008
Brown, J.H., Yang, Y., Reshetnikova, L., Gourinath, S., Süveges, D., Kardos, J., Hóbor, F, Reutzel, R., Nyitray, L., Cohen, C.: an unstable head-rod junction may promote folding into compact off-state conformation of regulated myosins, J. Mol. Biol. 375: 1434-43, 2008
Gáspári, Z., Pál, G., Perczel, A.: A redesigned genetic code for selective labeling in protein NMR, Bioessays 30: 772-780, 2008



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