Pharmaceutical Chemistry A Bioinformatics Approach for Homology Modeling and %LQGLQJ6LWH,GHQWL¿FDWLRQRI7ULRVHSKRVSKDWH,VRPHUDVHIURP Plasmodium falciparum 3D7 Ullah M, Hira J1, Ghosh T, Ishaque N, Absar N Department of Biochemistry and Biotechnology, University of Science and Technology Chittagong (USTC), Bangladesh, 1Department of Laboratory Medicine, Children’s and Women’s Health, Norwegian University of Science and Technology (NTNU), Trondheim, Norway Address for correspondence: Mr. M. Ullah, E-mail: [email protected] ABSTRACT Malaria is a major public health concern, and malarial parasites have developed resistance against the commonly DYDLODEOHGUXJV6RQRZDGD\VLWLVDPDMRUFRQFHUQWR¿QGRXWDQHZWDUJHWIRUGUXJWKHUDS\Plasmodium falciparum 'RQHRIWKHVWUDLQVRISODVPRGLXPVSHFLHVDOVRODFNVLQDIXQFWLRQDOWULFDUER[\OLFDFLGF\FOHDQGVROHO\GHSHQGHQW RQJO\FRO\VLVIRULWVHQHUJ\VXSSO\OLNHRWKHUSODVPRGLXPVSHFLHV$OWKRXJKHQ]\PHVRIPDODULDOSDUDVLWHKDYHEHHQ considered as potential antimalarial drug targets, a little is known about their structural biology. The tertiary structure of triose phosphate isomerase of P. falciparum 3D7 was determined by means of homology modeling through multiple DOLJQPHQWIROORZHGE\LQWHQVLYHRSWLPL]DWLRQDQGYDOLGDWLRQ7KHPRGHOLQJZDVGRQHE\6ZLVV0RGHO:RUNVSDFH 7KHREWDLQHGPRGHOZDVYHUL¿HGZLWKWKHVWUXFWXUHYDOLGDWLRQSURJUDPVVXFKDV352&+(&.9HULI\'DQG40($1 IRUUHOLDELOLW\7KHYHULI\'YDOXHRILQGLFDWHVWKDWWKHHQYLURQPHQWSUR¿OHRIWKHPRGHOLVJRRG$VHOIRSWLPL]HG SUHGLFWLRQPHWKRGZLWKDOLJQPHQWRU6230$LVHPSOR\HGIRUFDOFXODWLRQRIWKHVHFRQGDU\VWUXFWXUDOIHDWXUHVRI triose phosphate isomerase. The secondary structure indicates that the predicted 3D structure of triosephosphate isomerase of P. falciparum'FRQWDLQVDKHOL[UDQGRPFRLODQGH[WHQGHGVWUDQG$FWLYH VLWHGHWHUPLQDWLRQWKURXJK&$67SVXJJHVWVWKDWWKLVSURWHLQFDQEHXWLOL]HGDVDSRWHQWLDOGUXJWDUJHW+RZHYHU these will further be tested by wet lab studies for a targeted vaccine design against P. falciparum 3D7. Key words: Active site, homology modeling, Plasmodium falciparum 3D7 INTRODUCTION LWDPDMRUFDXVHRI KXPDQPRUELGLW\DQGPRUWDOLW\ ZRUOGZLGH>@7KHGLVHDVHLVFDXVHGE\WKHSDUDVLWHVRI 0DODULDDIÁLFWVPLOOLRQSHRSOHHDFK\HDUPDNLQJ the Plasmodium species (Plasmodium falciparum, Plasmodium vivax, Plasmodium ovale, and Plasmodium malariae DQG Access this article online LVWUDQVPLWWHGE\IHPDOHPRVTXLWRRI WKH$QRSKHOHV Quick Response Code: genus.>@ P. falciparum is the most virulent of the four Website: www.jyoungpharm.in species causing malaria and responsible for most PDODULDOGHDWKV7KHSDUWLFXODUYLUXOHQFHRI P. falciparum DOI: LVSDUWO\GXHWRWKHDELOLW\RI LQIHFWHGHU\WKURF\WHVWR 10.4103/0975-1483.104370 DGKHUHWRDYDULHW\RI KRVWUHFHSWRUVDQGDYRLGVSOHQLF clearance.>@ Because of the increased prevalence of P. Journal of Young Pharmacists Vol 4 / No 4 261 Ullah, et al+RPRORJ\PRGHOLQJDQGELQGLQJVLWHLGHQWL¿FDWLRQRIWULRVHSKRVSKDWHLVRPHUDVH falciparumVWUDLQVWREHUHVLVWDQWWRFXUUHQWFKHPRWKHUDS\ MATERIALS AND METHODS treatment and control of this disease are becoming SURJUHVVLYHO\PRUHGLIILFXOW>@7KHSKHQRW\SHVP. Computational methods falciparum'RULJLQDWHGIURPWKH1HWKHUODQGVUHSUHVHQW GLVWLQFWGUXJUHVLVWDQFHDQGRUVHQVLWLYLW\WRFRPPRQ 7KHFRPSXWDWLRQDOPHWKRGVRI 'PRGHOEXLOGLQJ LQYROYHGWHPSODWHVHOHFWLRQDOLJQPHQWRI WHPSODWHZLWK antimalarial chemotherapeutics such as chloroquine and the target, building of the model, and evolution of the sulfadoxine.>@7KHSDUDVLWHDOVRODFNVDIXQFWLRQDO VWUXFWXUH7KHVHTXHQFHRI WULRVHSKRVSKDWHLVRPHUDVH WULFDUER[\OLFDFLGF\FOH>@7KHDVH[XDOVWDJHRIWKH of P. falciparum'ZDVUHWULHYHGIURP1&%,GDWDEDVH SDUDVLWHUHVLGLQJLQWKHPDWXUHKXPDQHU\WKURF\WH $FFHVVLRQ,';3B VROHO\GHSHQGVRQJO\FRO\VLVIRULWV$GHQRVLQH WULSKRVSKDWH $73 UHTXLUHPHQWV7KHUHIRUHWKH Template selection and sequence alignment JO\FRO\WLFHQ]\PHVSURYLGHJRRGGUXJWDUJHWV>@7KH JOXFRVHFRQVXPSWLRQRI WKHLQIHFWHGHU\WKURF\WHV 7KH3'% %URRNKDYHQ3URWHLQ'DWDEDQN GDWDEDVH>@ZDV LQFUHDVHVE\WRIROGRYHUWKDWRI XQLQIHFWHG H[WHQVLYHO\VFUHHQHGXVLQJ%/$67 %DVLF/RFDO$OLJQPHQW HU\WKURF\WHV,QLQIHFWHGHU\WKURF\WHVWKHÁX[RI VHYHUDO 7RRO >@ server, developed and maintained in Adam JO\FRO\WLFHQ]\PHVDUHHOHYDWHG>@*O\FRO\WLFHQ]\PHVDUH *RG]LN·VODERUDWRU\DWWKH%XUQKDP,QVWLWXWHWRÀQGRXW EHOLHYHGWREHDVVRFLDWHGZLWKPHPEUDQHFRPSRQHQWV WKHUHODWHGKRPRORJVRI WKHTXHU\VHTXHQFH3'%GDWDEDVH IDFLOLWDWLQJFKDQQHOLQJRI WKHVXEVWUDWHEHWZHHQ ZDVVHDUFKHGE\36,²%/$67XVLQJDSURÀOHJHQHUDWHGIURP FRQVHFXWLYHJO\FRO\WLFHQ]\PHVGXULQJWULRVHSKRVSKDWH QUSURWHLQGDWDEDVHIRUSURWHLQVH[KLELWLQJVLPLODULW\WRWKH metabolism.>@*O\FRO\VLVLQWKHSDUDVLWHPLJKWEHLQKLELWHG XQNQRZQVWUXFWXUH HLWKHUE\GHVLJQLQJVSHFLÀFLQKLELWRUVIRUWKHHQ]\PHVRI Model building WKHSDWKZD\RUE\GLVUXSWLQJWKHPLFURDVVHPEO\RQWKH F\WRVNHOHWRQ7KUHHGLPHQVLRQDOVWUXFWXUHVRI JO\FRO\WLF 7KHSURJUDP&3+PRGHOVVHUYHU>@ZDVXVHGWREXLOG HQ]\PHVRI P. falciparum might be of undoubted value WKHPRGHODVD3'%ÀOHRI WULRVHSKRVSKDWHLVRPHUDVHRI LQGHVLJQLQJQHZVWUDWHJLHVIRUWKHUDSHXWLFLQWHUYHQWLRQ P. falciparum'DFFRUGLQJWRWKHKRPRORJ\PRGHOLQJ 7ULRVHSKRVSKDWHLVRPHUDVHLVDGLPHULFJO\FRO\WLFHQ]\PH PHWKRG7KHPRGHOZDVVXEPLWWHGWRWKH6ZLVV0RGHO ZKLFKFDWDO\]HVWKHLVRPHUL]DWLRQRI GJO\FHUDOGHK\GH :RUNVSDFH >@WRREWDLQWKH'VWUXFWXUH SKRVSKDWHWRGLK\GUR[\DFHWRQHSKRVSKDWH([WHQVLYH PXWDJHQHVLVH[SHULPHQWVRQWKH\HDVWSURWHLQVXJJHVW Evolution and validation of model WKDWWULRVHSKRVSKDWHLVRPHUDVHLVDQ´HYROXWLRQDULO\ SHUIHFWHQ]\PHµ>@7ULRVHSKRVSKDWHLVRPHUDVHDOVR 7KHVWHULRFKHPLFDOTXDOLW\RI WKHPRGHOVZDVYHULÀHG SOD\VDQLPSRUWDQWUROHLQJOXFRQHRJHQHVLVWKHKH[RVH ZLWKWKHSURJUDP352&+(&.>@ in order to select the PRQRSKRVSKDWHVKXQWDQGIDWW\DFLGELRV\QWKHVLV EHVWPRGHO7KH'SURÀOLQJRI WKHUHVLGXHZDVGRQHE\ >@ >@ It might be argued that the same triosephosphate 9HULI\'6WUXFWXUH(YDOXDWLRQ6HUYHU DQG40($1 LVRPHUDVHLVSUHVHQWLQKXPDQVWRRDQGKRZDGUXJFDQ Secondary structure and active site analysis EHWDUJHWHGLQVXFKDZD\WKDWLWGRHVQRWDIIHFWWKHKRVW 7KHWULRVHSKRVSKDWHLVRPHUDVHRI P. falciparumZKHQ 7KHVHFRQGDU\VWUXFWXUHHYROXWLRQZDVGRQHZLWKWKHKHOS FRPSDUHGWRKXPDQWULRVHSKRVSKDWHLVRPHUDVHVKRZV RI DVHOIRSWLPL]HGSUHGLFWLRQPHWKRGZLWKDOLJQPHQWRU >@ PDQ\LPSRUWDQWGLIIHUHQFHV 7KHUHKDVEHHQDUHFHQW 6230$>@$FWLYHVLWHDQDO\VLVZDVSHUIRUPHGZLWKWKH report that during malarial infection, patients mount KHOSRI &$67S>@IRUWKHPRGHOHGSURWHLQWRIXUWKHUZRUN DQDQWLERG\UHVSRQVHWRWULRVHSKRVSKDWHLVRPHUDVH RQLWVGRFNLQJVWXGLHV$FWLYHVLWHDQDO\VLVJLYHVXVDQLGHD UHVXOWLQJLQSURORQJHGKHPRO\WLFDQHPLD>@7KLVUDLVHV WRPDNHDJULGEHIRUHGRFNLQJ WKHSRVVLELOLW\WKDWWKHSURWHLQPD\EHFRPHORFDOL]HGRQ WKHHU\WKURF\WHPHPEUDQHDQGJHWWLQJSDUWO\H[SRVHG RESULTS Indeed, the triosephosphate isomerase from the parasite Schistosoma mansoni ZDVDOVRVKRZQWREHDVXUIDFHDQWLJHQ Template selection and sequence alignment and has been considered for vaccine development.>@ In this SDSHUZHKDYHUHSRUWHGWKH'VWUXFWXUHRI P. falciparum 7KHVWUXFWXUHRI WKHWULRVHSKRVSKDWHLVRPHUDVHRI P. VWUDLQ'DQGDOVRSUHGLFWHGLWVDFWLYHVLWHIRUDSRWHQWLDO falciparum'KDVEHHQFRQVWUXFWHGEDVHGRQWKHDYDLODELOLW\ GUXJWDUJHWE\FRPSXWHUVLPXODWLRQ of the template sequence of chain A, triosephosphate 262 Journal of Young Pharmacists Vol 4 / No 4 Ullah, et al+RPRORJ\PRGHOLQJDQGELQGLQJVLWHLGHQWL¿FDWLRQRIWULRVHSKRVSKDWHLVRPHUDVH isomerase of P. falciparum 3'%FRGH<'9B$ >@ Secondary structure and active site of triosephosphate isomerase 3D-model Analysis of secondary structure 7KH'VWUXFWXUHRI WULRVHSKRVSKDWHLVRPHUDVHRI P. falciparum 7KHVHFRQGDU\VWUXFWXUHDQDO\VLVLVXVHGWRNQRZ 'ZDVEXLOWE\KRPRORJ\PRGHOLQJEDVHGRQWKHWHPSODWH ZKHWKHUDJLYHQDPLQRDFLGOLHVLQDKHOL[VWUDQGRUFRLO ZKLFKLVREWDLQHGIURPSURWHLQGDWDEDQN>)LJXUH@ 6HFRQGDU\VWUXFWXUDOIHDWXUHVDVSUHGLFWHGXVLQJ6230$ DUHUHSUHVHQWHGLQ7DEOH7KHVHFRQGDU\VWUXFWXUHDUH Validation of model SUHGLFWHGE\XVLQJGHIDXOWSDUDPHWHUV ZLQGRZZLGWK VLPLODULW\WKUHVKROGDQGQXPEHURI VWDWHV Validation of the structure 7KHƶ and ƸGLVWULEXWLRQYDOXHRI WKH5DPDFKDQGUDQSORW Active site determination RI QRQJO\FLQHDQGQRQSUROLQHUHVLGXHVDUHVKRZQLQ 7KHDFWLYHVLWHRI WULRVHSKRVSKDWHLVRPHUDVHRI P. falciparum )LJXUHDQG7DEOH 'ZDVSUHGLFWHGE\XVLQJ&$67SVHUYHU>)LJXUHD@ )XUWKHULQWKLVVWXG\ZHKDYHDOVRUHSRUWHGWKHEHVWDFWLYH 9HULÀFDWLRQRI WKH'PRGHO VLWHDUHDRI WKHH[SHULPHQWDOHQ]\PHDVZHOODVWKHQXPEHU 7KHSURÀOHVFRUHDERYH]HURLQWKH9HULI\'JUDSKFRUUHVSRQG RI DPLQRDFLGLQYROYHGLQLW>)LJXUHE@ WRWKHDFFHSWDEOHHQYLURQPHQWRI WKHPRGHO>)LJXUH@7KH JOREDOTXDOLW\RI WKHPRGHOIRUWKHWDUJHWVHTXHQFHZDV HVWLPDWHGE\WKH40($1VFRUHEDVHGRQOLQHDUFRPELQDWLRQ DISCUSSION RI VL[VWUXFWXUDOGHVFULSWRUV>7DEOH@ZKLFKUHÁHFWVSUHGLFWHG 7KHKRPRORJ\PRGHOLQJDOJRULWKPKDVSURYHGWREHRQH PRGHOUHOLDELOLW\UDQJHVIURPWR>@7KHUDZ40($1 RI WKHPRVWUREXVWPRGHOLQJWRROVLQELRLQIRUPDWLFV7KH score and average ZVFRUHIRUWULRVHSKRVSKDWHLVRPHUDVHRI SURWHLQPRGHOEXLOWLQWKLVVWXG\E\XVLQJVWDQGDUGSURWRFRO P. falciparum'PRGHODUHVKRZQLQ)LJXUH LQGLFDWHVWKDWWKHHQ]\PHFRQWDLQVDKHOL[EVKHHWVDQG ORRSV7KHVWUXFWXUHPRGHOHGE\&3+PRGHOVVHUYHU Figure 1: 3D model of triose phosphate isomerase of P. falciparum 3D7 by comparative modeling (Swiss Model Workspace) Figure 2: Ramachandran plot of triose phosphate isomerase of P. Table 1: Ramachandran plot statistics of falciparum 3D7 obtained through the modeling tool triosephosphate isomerase of 3IDOFLSDUXP 3D7 Rama chandran plot statistics (%) Table 2: Z scores of individual component of QMEAN for Residues in the most favored regions [A, B, L] 204 90.3 WULRVHSKRVSKDWHLVRPHUDVH 3IDOFLSDUXP 3D7) models Residues in the additional allowed regions [a, b, l, p] 21 9.3 Scoring function term Z-score Residues in the generously allowed regions [a, b, l, p] 1 0.4 C_E interaction energy 0.16 Residues in the disallowed regions 0 0.0 All-atom pairwise energy í Number of non-glycine and non-proline residues 226 100 Solvation energy 1.72 Number of end-residues (excl. Gly and Pro) 2 Torsion angle energy í Number of glycine residues