Principal investigator:
Anastassios C. Papageorgiou, Ph.D., Adjunct Professor in Biochemistry and Structural Biology Turku Centre for
Biotechnology, BioCity, Tykistökatu 6A, FI-20521 Turku, Finland. Tel. +358-2-3338012, Fax +358-2-3338000.
Email: [email protected]
Biography:
Tassos Papageorgiou (b. 1962) obtained his Ph.D. from the University of Athens in 1992. He was a postdoctoral fellow at the University of Oxford and University of Bath (UK). In May 2000, he joined the Centre for Biotechnology as senior scientist in protein crystallography
Personnel:
Graduate students: Prathusha Dhavala, Teemu Haikarainen, Sachin Wakadkar
Undergraduate students: Simon Le Boulh, Pia Kinaret. Omid Mohammadi, Bishwa Subedi, Kristian Wecström
Description of the projects:
We use X-crystallography, molecular biology and biophysical techniques to study the structure and function of biological molecules. One of our major projects has been the Dps family of proteins that are widely spread among procaryotes and responsible for protection against oxidative stress due to their ability to oxidize and store iron. Although Dps proteins are structurally similar to ferritins, they form a spherical shell of 12 subunits instead of 24 and have a different ferroxidase center compared to that of ferritins. The crystal structures of Dps-like peroxide resistance protein (Dpr) from the pathogenic bacterium Streptococcus suis in the iron-free, the iron-bound and zinc-bound form have been recently determined. In addition, EXAFS experiments performed at EMBL Hamburg have provided detailed information of the geometry of the iron core and showed a ferrihydrite-like structure. Based on our recent results, a number of mutants have been generated to study the iron core formation using X-ray crystallography, microcalorimetry, EXAFS, magnetization and Mössbauer spectroscopy techniques. The structure of Dpr from Streptococcus pyogenes was determined in 2009 and structural studies on Helicobacter pylori neutrophil- activating protein (known as HP-NAP) were initiated to identify potential ligand binding sites.
Studies on oxidative stress protection and detoxification mechanisms have been extended by determining high-resolution crystal structures of a tau family glutathione transferase (GST) from Glycine max in free form and in the presence of a substrate analogue. Importantly, the crystal structures revealed a novel site on the surface of the protein that may be utilised for storage and/ or transport of dangerous compounds for detoxification. Docking calculations were carried out to study the binding of diphenylether herbicides in the active site. Work is currently underway on chimeric GSTs or mutants created through directed evolution approaches to produce new GSTs with altered specificity for new applications in agriculture and biomedicine. Crystals of human GST-A1 have been grown in our lab for use in structure-assisted drug design efforts. In addition, the structure of a novel glutathione transferase Lund, R., Chen, Z., Scheinin, J. and Lahesmaa, R. (2004) Early
target genes of IL-12/STAT4 signaling in T helper cells. J. Immunol. 172: 6775-6782.
Lund R*, Pykäläinen M*, Naumanen T, Dixon C, Chen Z, Ahlfors H, Tuomela S, Tahvanainen J, Scheinin J, Henttinen T, Rasool O, Lahesmaa R. (2007) Genome wide identification of Novel Genes Involved in Early Th1 and Th2 Cell Differentiation J. Immunol. 178:3648-60.
Moulder R*, Lönnberg T*, Filén J-J, Elo L, Rainio E, Corthals G, Oresic M, Nyman TA, Aittokallio T, Lahesmaa R (2010) (*equal contribution). Quantitative Proteomics Analysis of the Nuclear Fraction of Human CD4+ Cells in the Early Phases of IL-4 Induced Th2 Differentiation. Mol Cell Proteomics. 9:1937-53.
Närvä E, Autio R, Rahkonen N, Kong L, Harrison N, Kitsberg D, Borghese L, Itskovitz-Eldor J, Rasool O, Dvorak P, Hovatta O, Otonkoski T, Tuuri T, Cui W, Brüstle O, Baker D, Maltby E, Moore HD, Benvenisty N, Andrews PW, Yli-Harja O & Lahesmaa R. (2010) High resolution genome wide DNA analysis on a large panel of Human Embryonic Stem Cell lines reveals novel genomic changes associated with culture and affecting gene expression.
Nat Biotechnol. 28:371-7.
Oresic M, Simell S*, Sysi-Aho M*, Näntö-Salonen K*, Seppänen- Laakso T*, Parikka V*, Katajamaa M*, Hekkala A, Mattila I, Keskinen P, Yetukuri L, Reinikainen A, Lähde J, Suortti T, Hakalax J, Simell T, * Equal contribution. J Exp Med. 205:2975-84.
Rautajoki, K., Marttila, E., Nyman, T., Lahesmaa, R. (2007) Interleukin-4 inhibits caspase-3 by regulating several proteins in the Fas pathway during initial stages of human T helper 2 cell differentiation. Mol. Cell Proteomics. 6: 238-251.
Tahvanainen J, Kallonen T, Lähteenmäki H, Heiskanen KM, Westermarck J, Rao KV, Lahesmaa R. (2008)
Blood. 113:1268-77.
Tuomela S, Rautajoki KJ, Moulder R, Nyman TA, Lahesmaa R. (2009) Identification of novel Stat6 regulated proteins in IL-4-treated mouse lymphocytes. Proteomics. 9:1087-98.
was determined by the SAD method using the anomalous signal of Br. The overall fold and the geometry of the active site suggest a new member of the glutathione transferase superfamily. Knock-out and microarray experiments are currently in progress to provide functional insights.
In the theme of enzyme function and stability, we continued our work on PhaZ7, an extracellular depolymerase involved in the degradation of poly(R)-hydroxyalkanoates, a group of thermoplastic polyesters considered as biodegradable substitutes for non- degradable plastics. The crystal structure of PhaZ7 depolymerase at atomic (1.2) Å resolution in the presence of the serine protease inhibitor PMSF was determined. A molecule of SO2 was found covalently bound at the active site of the enzyme, suggesting a preformed catalytic triad. Several mutants have been generated by our collaborators and characterized. Crystal structure determination is currently underway. The structure of Erwinia carotovora asparaginase at 1.4 Å, the second highest resolution for an L-asparaginase was determined. Asparaginases are widely- used enzymes in leukemia treatment for the last 30 years. However, asparaginases from Erwinia chrysanthemi and E. coli cause severe side-effects. Protein engineering efforts have been initiated for the Erwinia carotovora enzyme based on its 3D-structure to produce chimeric forms with reduced glutaminase activity (the main reason of the side-effects) and high specificity for L-asparagine. Work on the Atu (acyclic terpene utilization) catabolic pathway found in P. Aeruginosa has been initiated using a combination of X-ray crystallography, biophysics, molecular biology, homology modelling, computational and bioinformatics tools. Atu enzymes are involved in the metabolisn of acyclic terpenes that possess a great potential in biotechnology, for example in the food, drink and pharmaceutical industry.
Selected publications:
Wakadkar, S., Zhang,L.Q., Li, D.-C., Haikarainen, T., Dhavala, P. & Papageorgiou, A.C. (2010). Expression, purification and crystallization of Chetomium thermophilum Cu, Zn superoxide dismutase. Acta Cryst F (in press).
Haikarainen, T., Tsou, C.C., Wu, J.J. & Papageorgiou, A.C. (2010). Structural characterization and biological implications of di-zink binding in the ferroxidase center of Strepococcus pyogenes Dpr.
Bichem. Biophys. Res. Comm. 398, 361-365.
Haikarainen, T. & Papageorgiou, A.C. (2010). Dps-like proteins: Structural and functional insights into a versatile protein family. Cell.
Mol. Life Sci. 67, 341-351.
Axarli, I., Georgiadou, C., Dhavala, P., Papageorgiou, A.C. & Labrou, N. (2010). Investigation of the role of conserved residues Ser13, Asn48 and Pro49 in the catalytic mechanism of the tau class glutathione transferase from Glycine max. Bioch. Biophys.
Acta 1804, 662-667.
Labrou, N., Papageorgiou, A.C. & Avramis, V.I. (2010). Structure- function relationships and clinical applications of L-asparaginases.
Curr. Med. Chem. 17, 2183-2195.
Wakadkar, S., Hermawan, S., Jendrossek, D. & Papageorgiou, A.C. (2010). The crystal structure of PhaZ7 at atomic (1.2 Å) resolution reveals details of the active site and suggests a substrate-binding mode. Acta Cryst. F 66, 648-654.
Melissis, S.C., Papageorgiou, A.C., Labrou, N.E & Clonis, Y.D. (2010). Purification of moloney murine leukemia virus reverse transcriptase lacking RNase activity (M-MLVH-RT) on a 9-aminoethyladenine- [1,6-diamine-hexane]-triazine selected from a combinatorial library of dNTP-mimetic ligands. J. Chromatogr. Sci. 48, 496-502. Haikarainen, T., Tsou, C.C., Wu, J.J. & Papageorgiou, A.C. (2010). Crystal structures of Streptococcus pyogenes Dpr reveal a dodecameric iron-binding protein with a ferroxidase site. J. Biol.
Inorg. Chem. 15, 183-194.
Axarli, I. Dhavala, P., Papageorgiou, A.C. & Labrou, N.E. (2009). Crystallographic and functional characterization of the fluorodifen- inducible glutathione transferase from Glycine max reveals an active site topography suited for diphenylether herbicides and a novel L-site. J. Mol. Biol. 385, 984-1002.
Axarli, I. Dhavala, P., Papageorgiou, A.C. & Labrou, N.E. (2009). Crystal structrure of Glycine max glutathione transferase in complex with glutathione: investigation of the induced-fit mechanism operating by the tau class glutathione transferases. Biochem. J. 422, 247-256.
Mitsiki, E., Papageorgiou, A. C., Iyer, S., Thiyagarajan, N., Prior, S. H., Sleep, D., Finnis, C. & Acharya, K. R. (2009). Structures of native human thymidine phosphorylase and in complex with 5-iodouracil.
Biochem. Biophys. Res. Commun. 386, 666-670.
Dhavala, P. & Papageorgiou, A.C. (2009). The crystal structure of Helicobacter pylori L-asparaginase at 1.4 Å resolution. Acta
Crystallogr. D 65, 1253-1261.
Havukainen, H., Haataja, S., Kauko, A., Pulliainen, A.T., Salminen, A., Haikarainen, T., Finne, J. & Papageorgiou, A.C. (2008). Structural basis of zinc- and terbium-mediated inhibition of ferroxidase activity in Dps ferritin-like proteins. Protein Sci. 17, 1513-1521
Papageorgiou, A.C., Posypanova, G.A., Andersson, C.A., Sokolov, N.N & Krasotkina, J. (2008) Structural and functional insights into Erwinia carotovora L-asparaginase. FEBS J. 275, 4306-4316 Dhavala, P., Krasotkina, J., Dubreuil, C. & Papageorgiou, A.C. (2008). Expression, purification and crystallization of Helicobacter pylori L-asparaginase. Acta Crystallogr Sect F Struct Biol Cryst
Commun. 64, 740-742
Papageorgiou, A.C., Hermawan, S., Singh C.B. & Jendrossek, D. (2008) Structural basis of poly(3-hydroxybutyrate) hydrolysis by PhaZ7 depolymerase from Paucimonas lemoignei. J. Mol. Biol. 382, 1184-1194
Saarinen S., Kato, H., Uchiyama, T., Miyoshi-Akiyama, T. & Papageorgiou, A.C. (2007). Crystal structure of Streptococcus dysgalactiae-derived mitogen reveals a zinc-binding site and alterations in TcR binding. J. Mol. Biol. 373, 1089-1097
Weckström, K. & Papageorgiou, A.C. (2007). Lower consolute boundaries of the nonionic surfactant C(8)E(5) in aqueous alkali halide solutions: An approach to reproduce the effects of alkali halides on the cloud-point temperature. J Colloid Interface Sci. 310, 151-162
Zhao, J., Hayashi, T., Saarinen, S., Papageorgiou, A.C., Kato, H., Imanishi, K., Kirikae, T., Abe, R., Uchiyama, T. & Miyoshi- Akiyama, T. (2007). Cloning, expression and characterization of the superantigen streptococcal pyrogenic exotoxin-G from Streptococcus dysgalactiae. Inf. Immun. 75, 1721-1729
Kauko, A., Pulliainen, A.T., Haataja, S., Meyer-Klaucke, W., Finne, J. & Papageorgiou, A.C. (2006). Iron incorporation in Streptococcus suis Dps-like peroxide resistance protein Dpr requires mobility in the ferroxidase center and leads to the formation of a ferrihydrite- like core. J. Mol. Biol. 364: 97-109
Papageorgiou, A.C., Saarinen, S., Ramirez-Bartutis, R., Kato, H., Uchiyama, T., Kirikae, T. & Miyoshi-Akiyama, T. (2006). Expression, purification and crystallisation of Streptococcus dysgalactiae- derived mitogen. Acta Crystallogr. F. 62: 242-244
Kapetaniou, E.G., Thanassoulas, A., Dubnovitsky, A.P., Nounesis, G. & Papageorgiou, A.C. (2006). The effect of pH on the structure and stability of Bacillus circulans ssp. alkalophilus phosphoserine aminotransferase: Thermodynamic and crystallographic studies.
Proteins: Struct. Funct. Bioinform. 63: 742-753
Pulliainen, A.T., Kauko, A., Haataja, S., Papageorgiou, A.C. & Finne, J. (2005). Dpr/Dps miniferritin: Insights into the mechanism of iron incorporation and evidence for a central role in cellular iron homeostasis in Streptococcus suis. Mol. Microbiol. 57, 1086- 1100.
Kapetaniou, E.G, Braaz, R., Jendrossek, D. & Papageorgiou, A.C. (2005). Crystallization and preliminary X-ray analysis of a novel thermoalkalophilic depolymerase (PhaZ7) from Paucimonas lemoignei. Acta Crystallogr. F 61: 479-481
Wikman, L.E.K., Krasotkina, J., Kuchumova, A., Sokolov, N.N. & Papageorgiou, A.C. (2005). Crystallization and preliminary crystallographic analysis of L-asparaginase from Erwinia carotovora.
Acta Crystallogr. F 61: 407-409
Dubnovitsky,. A.P., Ravelli, R.B.G., Popov, A.N. & Papageorgiou, A.C. (2005). Strain relief at the active site of phosphoserine aminotransferase induced by radiation damage. Protein Sci. 14: 1498-1507
Mialon A., Sankinen, M., Söderström, H., Junttila, T.T., Holmström, T., Koivusalo, R., Papageorgiou, A.C., Johnson, R.S., Hietanen, S., Elenius, K. & Westermarck, J. (2005). DNA topoisomerase I is a co- factor for c-Jun in the regulation of EGFR expression and cancer cell proliferation. Mol. Cell. Biol. 25: 5040-5051
Dubnovitsky, A.P., Kapetaniou, E.G. & Papageorgiou, A.C. (2005). Enzyme adaptation to alkaline pH: Atomic resolution (1.08 Å) structure of phosphoserine aminotransferase from Bacillus alkalophilus. Protein Sci. 14: 97-110
From left to right, sitting: Bishwa Subedi, Teemu Haikarainen, back row: Tassos Papageorgiou, Sachin Wakadkar, Prathusha Dhavala.