Sains Malaysiana 45(12)(2016): 1947–1957

http://dx.doi.org/10.17576/jsm-2016-4512-20

 

Pencirian Molekul Glikogen Sintase Kinase-3 dari Eimeria tenella

(Molecular Characterisation of Glycogen Synthase Kinase-3 from Eimeria tenella)

 

PING-PING YAO, MOHD FIRDAUS RAIH, HASIDAH MOHD SIDEK, NOOR EMBI

 & KIEW-LIAN WAN*

 

Pusat Pengajian Biosains & Bioteknologi, Fakulti Sains & Teknologi, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor Darul Ehsan, Malaysia

 

Received: 14 April 2016/Accepted: 26 October 2016

 

ABSTRAK

Penemuan sasaran dadah antikoksidia baharu merupakan antara usaha yang diperlukan untuk mengawal penyakit koksidiosis ayam yang disebabkan oleh spesies Eimeria. Dalam kajian ini, serpihan yang mengekodkan glikogen sintase kinase-3 (GSK-3) Eimeria tenella putatif telah diamplifikasi daripada cDNA E. tenella. Hasil pemadanan homologi menunjukkan jujukan GSK-3 E. tenella yang terjana mempunyai padanan yang tinggi dengan jujukan GSK-3 organisma lain. Domain terpulihara GSK-3 dan residu yang penting untuk aktiviti GSK-3 juga diramalkan hadir dalam jujukan GSK-3 E. tenella. Analisis struktur sekunder serta pemodelan homologi menunjukkan pembahagian struktur protein kepada domain bebenang beta pada hujung N dan domain heliks alfa pada hujung C, yang merupakan ciri enzim GSK-3. Kesemua hasil analisis ini menyokong bahawa jujukan yang dikaji mengekodkan protein GSK-3 dalam E. tenella. Walaupun darjah keterpuliharaan adalah tinggi, namun terdapat perbezaan yang bermakna diperhatikan antara GSK-3 E. tenella dan perumahnya. Residu Ser 9 yang dilaporkan penting untuk perencatan aktiviti GSK-3 didapati tidak terpulihara dalam GSK-3 E. tenella. Memandangkan Ser 9 merupakan tapak pemfosfatan bagi GSK-3β dalam haiwan vertebrata, ketiadaan residu ini dalam jujukan GSK-3 E. tenella mencadangkan bahawa pengawalaturan GSK-3 E. tenella melibatkan tapak pemfosfatan dan mekanisme yang berbeza. Tambahan pula, hasil analisis filogenetik menunjukkan bahawa GSK-3 E. tenella mempunyai pertalian yang rapat dengan protein GSK-3 tumbuh-tumbuhan. Analisis superposisi GSK-3 E. tenella dengan GSK-3β Homo sapiens pula menunjukkan bahawa perencat GSK-3 mampu berinteraksi dengan protein GSK-3 E. tenella. Keputusan kajian ini mencadangkan bahawa GSK-3 E. tenella mempunyai potensi untuk diperkembangkan sebagai sasaran dadah antikoksidia.

 

Kata kunci: Koksidiosis; parasit protozoa; sasaran dadah anti-koksidia

 

ABSTRACT

The discovery of new anticoccidial drug targets is amongst the necessary efforts needed to control chicken coccidiosis caused by Eimeria species. In this study, the fragment coding for the putative Eimeria tenella glycogen synthase kinase-3 (GSK-3) was amplified from the cDNA of E. tenella. Homology search showed that generated E. tenella GSK-3 sequence has high similarities with GSK-3 sequences from other organisms. The conserved domains of GSK-3 and residues important for the GSK-3 activity were also predicted within the E. tenella GSK-3. Secondary structure analysis and homology modelling predicted that the protein structure is divided into a beta strand domain at the N terminal and an alpha helix domain at terminal C, which are characteristics of GSK-3 enzymes. These results supported the E. tenella GSK-3 codes for the GSK-3 protein in E. tenella. Although the degree of conservation is high, significant differences were observed between GSK-3 of E. tenella and its host. The Ser 9 residue reported to be important for the inhibition of the GSK-3 activity was not conserved within the E. tenella GSK-3. Considering that Ser 9 is a phosphorylation site in GSK-3β of vertebrates, the absence of this residue in the E. tenella GSK-3 sequence suggests that the regulation of E. tenella GSK-3 involves a different phosphorylation site and mechanism. Phylogenetic analysis suggests that E. tenella GSK-3 has a closer relationship to plant GSK-3. Superposition analysis between E. tenella GSK-3 and Homo sapiens GSK-3β predicted that E. tenella GSK-3 is able to interact with a GSK-3 inhibitor. Taken together, these results suggested that the E. tenella GSK-3 has the potential to be developed into an anticoccidial drug target.

 

Keywords: Anti-coccidial drug target; coccidiosis; protozoan parasite

REFERENCES

Ali, A., Hoeflich, K.P. & Woodgett, J.R. 2001. Glycogen synthase kinase-3: Properties, functions and regulations. Chemical Reviews 101: 2527-2540.

Allen, P.C. & Fetterer, R.H. 2002. Recent advances in biology and immunobiology of Eimeria species and in diagnosis and control of infection with these coccidian parasites of poultry. Clinical Microbiology Reviews 15(1): 58-65.

Altschul, S.F., Madden, T.L., Schaffer, A.A., Zhang, J., Zhang, Z., Miller, W. & Lipman, D.J. 1997. Gapped BLAST and PSI-BLAST: A new generation of protein database search programs. Nucleic Acids Research 25(17): 3389-3402.

Azizan, S., Wan, K.L. & Mohd-Adnan, A. 2014. Molecular characterisation and expression analysis of cathepsin D from the Asian seabass Lates calcarifer. Sains Malaysiana 43(8): 1139-1148.

Biasini, M., Bienert, S., Waterhouse, A., Arnold, K., Studer, G., Schmidt, T., Kiefer, F., Cassarino, T.G., Bertoni, M., Bordoli, L. & Schwede T. 2014. Swiss-Model: Modelling protein tertiary and quaternary structure usinge evolutionary information. Nucleic Acids Research 42(Web Server issue): 252-258.

Blake, D.P. & Tomley, F.M. 2014. Securing poultry production from the ever-present Eimeria challenge. Trends in Parasitology 30: 12-19.

Bonfield, J.K., Smith, K.F. & Staden, R. 1996. A new DNA sequence assembly program. Nucleic Acids Research 23: 4992-4999.

Chapman, H.D. 1997. Biochemical, genetic and applied aspects of drug resistance in Eimeria parasites of the fowl. Avian Pathology 26(2): 221-244.

Chapman, H.D., Jeffers, T.K. & Williams, R.B. 2010. Forty years of monensin for the control of coccidiosis in poultry. Poultry Science 89(9): 1788-1801.

Cross, D.A., Alessi, D.R., Cohen, P., Andjelkovich, M. & Hemmings, B.A. 1995. Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature 378(6559): 785-789.

Dajani, R., Fraser, E., Roe, S.M., Yeo, M., Good, V.M., Thompson, V., Dale, T.C. & Pearl, L.H. 2003. Structural basis for recruitment of glycogen synthase kinase 3 beta to the axin-APC scaffold complex. EMBO Journal 22(3): 494- 501.

Droucheau, E., Primot, A., Thomas, V., Mattei, D., Knockaert, M., Richardson, C., Sallicandro, P., Alano, P., Jafarshad, A., Baratte, B., Kunick, C., Parzy, D., Pearl, L., Doerig, C. & Meijer, L. 2004. Plasmodium falciparum glycogen synthase kinase-3: molecular model, expression, intracellular localisation and selective inhibitors. Biochimica et Biophysica Acta 1697(1-2): 181-196.

Embi, N., Rylatt, D.B. & Cohen, P. 1980. Glycogen synthase kinase-3 from rabbit skeletal muscle-separation from cyclic-AMP-dependent protein kinase and phosphorylase kinase. European Journal of Biochemistry 107(2): 519-527.

Felsenstein, J. 1989. PHYLIP - Phylogeny Inference Package (Version 3.2). Cladistics 5: 164-166.

Frame, S. & Cohen, P. 2001. GSK3 takes centre stage more than 20 years after its discovery. Biochemical Journal 359: 1-16.

Hall, T.A. 1999. BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/ NT. Nucleic Acids Symposium Series 41: 95-98.

Hanks, S.K. & Quinn, A.M. 1991. Protein kinase catalytic domain sequence database: Identification of conserved features of primary structure and classification of family members. Methods in Enzymology 200: 38-62.

Janouškovec, J., Horák, A., Oborník, M., Lukeš, J. & Keeling, P.J. 2010. A common red algal origin of the apicomplexan, dinoflagellate and heterokont plastids. Proceedings of the National Academy of Sciences U.S.A. 107(24): 10949- 10954.

Johan, N., Jangi, M.S. & Wan, K.L. 2011. Pemencilan dan pencirian populasi Eimeria tenella daripada ayam hutan tempatan. Sains Malaysiana 38(6): 939-945.

Jonak, C. & Hirt, H. 2002. Glycogen synthase kinase 3/ SHAGGY-like kinases in plants: An emerging family with novel functions. Trends in Plant Science 7(10): 457-461.

Jones, D.T. 1999. Protein secondary structure prediction based on position-specific scoring matrices. Journal of Molecular Biology 292(2): 195-202.

Jope, R.S. & Johnson, G.V. 2004. The glamour and gloom of glycogen synthase kinase-3. Trends in Biochemical Sciences 29(2): 95-102.

Kohler, S., Delwiche, C.F., Denny, P.W., Tilney, L.G., Webster, P., Wilson, R.J., Palmer, J.D. & Roos, D.S. 1997. A plastid of probable green algal origin in Apicomplexan parasites. Science 275(5305): 1485-1489.

Kuo, C.H., Wares, J.P. & Kissinger, J.C. 2008. The Apicomplexan-whole genome phylogeny: an analysis of incongruence among gene trees. Molecular Biology and Evolution 25(12): 2689-2698.

Labbé, M., Péroval, M., Bourdieu, C., Girard-Misguich, F. & Péry, P. 2006. Eimeria tenella enolase and pyruvate kinase: A likely role in glycolysis and in others functions. International Journal for Parasitology 36(14): 1443-1452.

Larkin, M.A., Blackshields, G., Brown, N.P., Chenna, R., McGettigan, P.A., McWilliam, H., Valentin, F., Wallace, I.M., Wilm, A., Lopez, R., Thompson, J.D., Gibson, T.J. & Higgins, D.G. 2007. Clustal W and Clustal X version 2.0. Bioinformatics 23(21): 2947-2948.

Lau, A.O.T., McElwain, T.F., Brayton, K.A., Knowles, D.P. & Roalson, E.H. 2009. Babesia bovis: A comprehensive phylogenetic analysis of plastid-encoded genes supports green algal origin of apicoplasts. Experimental Parasitology 123(3): 236-243.

Lee, J.H., Wan, K.L., Mohd-Adnan, A. & Gabaldón, T. 2012. Evolution of the ferritin family in vertebrates. Trends in Evolutionary Biology 4: e3.

Loo, S.S., Blake, D.P., Mohd-Adnan, A., Mohamed, R. & Wan, K.L. 2010. Eimeria tenella glucose-6-phosphate isomerase: Molecular characterization and assessment as a target for anti-coccidial control. Parasitology 137: 1169-1177.

Meijer, L., Skaltsounis, A.L., Magiatis, P., Polychronopoulos, P., Knockaert, M., Leost, M., Ryan, X.P., Vonica, C.A., Brivanlou, A., Dajani, R., Crovace, C., Tarricone, C., Musacchio, A., Roe, S.M., Pearl, L. & Greengard, P. 2003. GSK-3 selective inhibitors derived from Tyrian Purple Indirubins. Chemistry & Biology 10(12): 1255-1266.

Ojo, K.K., Gillespie, J.R., Riechers, A.J., Napuli, A.J., Verlinde, C.L., Buckner, F.S., Gelb, M.H., Domostoj, M.M., Wells, S.J., Scheer, A., Wells, T.N. & Van Voorhis, W.C. 2008. Glycogen synthase kinase 3 is a potential drug target for African trypanosomiasis therapy. Antimicrobial Agents and Chemotherapy 52(10): 3710-3717.

Osolodkin, D.I., Zakharevich, N.V., Palyulin, V.A., Danilenko, V.N. & Zefirov, N.S. 2011. Bioinformatic analysis of glycogen synthase kinase 3: Human versus parasite kinases. Parasitology 138(6): 725-735.

Pettersen, E.F., Goddard, T.D., Huang, C.C., Couch, G.S., Greenblatt, D.M., Meng, E.C. & Ferrin, T.E. 2004. UCSF Chimera: A visualization system for exploratory research and analysis. Journal of Computational Chemistry 25(13): 1605-1612.

Qin, C.L., Tang, J. & Kim, K. 1998. Cloning and in vitro expression of TPK3, a Toxoplasma gondii homologue of shaggy/glycogen synthase kinase-3 kinases. Molecular and Biochemical Parasitology 93(2): 273-283.

Reid, A.J., Blake, D.P., Ansari, H.R., Billington, K., Browne, H.P., Bryant, J., Dunn, M., Hung, S.S., Kawahara, F., Miranda- Saavedra, D., Malas, T.B., Mourier, T., Naghra, H., Nair, M., Otto, T.D., Rawlings, N.D., Rivailler, P., Sanchez-Flores, A., Sanders, M., Subramaniam, C., Tay, Y.L., Woo, Y., Wu, X., Barrell, B., Dear, P.H., Doerig, C., Gruber, A., Ivens, A.C., Parkinson, J., Rajandream, M.A., Shirley, M.W., Wan, K.L., Berriman, M., Tomley, F.M. & Pain, A. 2014. Genomic analysis of the causative agents of coccidiosis in domestic chickens. Genome Research 24: 1676-1685.

Shirley, M.W., Smith, A.L. & Tomley, F.M. 2005. The biology of avian Eimeria with an emphasis on their control by vaccination. Advances in Parasitology 60: 285-330.

Sievers, F., Wilm, A., Dineen, D., Gibson, T.J., Karplus, K., Li, W., Lopez, R., McWilliam, H., Remmert, M., Soding, J., Thompson, J.D. & Higgins, D.G. 2011. Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega. Molecular System Biology 7: 539.

Soon, P.E., Tomley, F.M., Jangi, M.S. & Wan, K.L. 2006. Pengenalpastian protein membran putatif dalam sporozoit Eimeria tenella melalui penyaringan imuno. Sains Malaysiana 35(2): 23-28.

Talavera, G. & Castresana, J. 2007. Improvement of phylogenies after removing divergent and ambiguously aligned blocks from protein sequence alignments. Systematic Biology 56: 564-577.

ter Haar, E., Coll, J.T., Austen, D.A., Hsiao, H.M., Swenson, L. & Jain, J. 2001. Structure of GSK3 beta reveals a primed phosphorylation mechanism. Nature Structural Biology 8(7): 593-596.

Van de Peer, Y., Baldauf, S.L., Doolittle, W.F. & Meyer, A. 2000. An updated and comprehensive rRNA phylogeny of (crown) eukaryotes based on rate-calibrated evolutionary distances. Journal of Molecular Evolution 51(2000): 565-576.

Wang, Q.M., Fiol, C.J., DePaoli-Roach, A.A. & Roach, P.J. 1994. Glycogen synthase kinase-3 beta is a dual specificity kinase differentially regulated by tyrosine and serine/threonine phosphorylation. Journal of Biological Chemistry 269(20): 14566-14574.

Williams, R.B. 2002. Fifty years of anticoccidial vaccines for poultry (1952-2002). Avian Diseases 46(4): 775-802.

Xiao, J.F., Li, Z.S., Sun, M., Zhang, Y. & Sun, C.C. 2004. Homology modeling and molecular dynamics study of GSK3/ SHAGGY-like kinase. Computational Biology and Chemistry 28(3): 179-188.

Xingi, E., Smirlis, D., Myrianthopoulos, V., Magiatis, P., Grant, K.M., Meijer, L., Mikros, E., Skaltsounis, A.L. & Soteriadou, K. 2009. 6-Br-5methylindirubin-3’oxime (5-Me-6-BIO) targeting the leishmanial glycogen synthase kinase-3 (GSK- 3) short form affects cell-cycle progression and induces apoptosis-like death: Exploitation of GSK-3 for treating leishmaniasis. International Journal for Parasitology 39(12): 1289-1303.

 

 

*Corresponding author; email: klwan@ukm.edu.my