The structural insight of class III of polyhydroxyalkanoate synthase from Bacillus sp. PSA10 as revealed by in silico analysis

Listia Pradani; Muhammad Saifur Rohman; Sebastian Margino

doi:10.22146/ijbiotech.53717

The structural insight of class III of polyhydroxyalkanoate synthase from Bacillus sp. PSA10 as revealed by in silico analysis

https://doi.org/10.22146/ijbiotech.53717

Listia Pradani⁽¹⁾, Muhammad Saifur Rohman^(2*), Sebastian Margino⁽³⁾

(1) Graduate Program in Biotechnology, Graduate School of Universitas Gadjah Mada, Jl. Teknika Utara, Yogyakarta, 55281, Indonesia
(2) Graduate Program in Biotechnology, Graduate School of Universitas Gadjah Mada, Jl. Teknika Utara, Yogyakarta, 55281, Indonesia; Laboratory of Agricultural Microbiology, Faculty of Agriculture, Universitas Gadjah Mada, Yogyakarta, Indonesia
(3) Laboratory of Agricultural Microbiology, Faculty of Agriculture, Universitas Gadjah Mada, Yogyakarta, Indonesia
(*) Corresponding Author

Abstract

PhaC synthase is an enzyme responsible for PHA polymerization. In this work, the catalytic mechanism class III of PhaC synthase from Bacillus sp. PSA10 (BacPhaCSynt) was reported through in silico modelling approach based on the primary sequence of the PhaC synthase. The open reading frame BacPhaCSynt has been successfully isolated, cloned and overexpressed the recombinant protein in Escherichia coli BL21(DE3). To know the global architecture and catalytic mechanism, the structural prediction of BacPhaCSynt has been carried out by using MODELLER. The recombinant BacPhaCSynt exhibited monomeric molecular weight (MW) of 43.6 kDa, when it was analyzed on 12% SDS‐PAGE gel. Based on the structural prediction, BacPhaCSynt exhibited global architecture of α/β hydrolase fold, with the root mean square deviation (r.m.s.d) value of 0.94Å. The catalytic residues composition of BacPhaCSynt consists of C151, D307, and H336, but the H336 and D307 residues of the model have been distorted 62.8o and 175.2o from the corresponding residues of the template. Since the D307 is quite a distance from the H336, it might act as a general base for the activation of ‐OH group of the substrate. The results strongly suggested that the mode of action of BacPhaCSynt obeyed the covalent catalysis mechanism.

Keywords

polyhydroxyalkanoate (PHA); class III of PhaC synthase; α/β hydrolase; Bacillus sp. PSA10; MODELLER

Full Text:

PDF

References

Berger E, Ramsay BA, Ramsay JA, Chavarie C, Braunegg G. 1989. PHB recovery by hypochlorite digestion of nonPHB biomass. Biotechnol Tech. 3(4):227–232. doi:10.1007/BF01876053.

Braunegg G, Bona R, Koller M. 2004. Sustainable polymer production. Polym Plast Technol Eng. 43(6):1779–1793. doi:10.1081/PPT200040130.

Chek MF, Hiroe A, Hakoshima T, Sudesh K, Taguchi S. 2019. PHA synthase (PhaC): interpreting the functions of bioplasticproducing enzyme from a structural perspective. Appl. Microbiol. Biotechnol. 103(3):1131–1141. doi:10.1007/s0025301895388.

Chek MF, Kim SY, Mori T, Arsad H, Samian MR, Sudesh K, Hakoshima T. 2017. Structure of polyhydroxyalkanoate (PHA) synthase PhaC from Chromobacterium sp. USM2, producing biodegradable plastics. Sci Rep. 7(1):1–15. doi:10.1038/s4159801705509 4.

Chen GQ, Patel MK. 2012. Plastics derived from biological sources: present and future: a technical and environmental review. Chem Rev. 112(4):2082–2099. doi:10.1021/cr200162d.

Chovancová E, Pavelka A, Benes P, Strnad O, Brezovsky J, Kozlikova B, Gora A, Sustr V, Klvana M, Medek P, Biedermannová L, Sochor J, Damborský J. 2012. CAVER 3.0: a tool for the analysis of transport pathways in dynamic protein structures. PLoS Comput Biol. 8(10):e1002708. doi:10.1371/journal.pcbi.1002708.

Hooft RW, Vriend G, Sander C, Abola EE. 1996. Errors in protein structure. Nature 381:272–272. doi:10.1038/381272a0.

Hu WF, Sin SN, Chua H, Yu PHF. 2005. Synthesis of polyhydroxyalkanoate (PHA) from excess activated sludge under various oxidationreduction potentials (ORP) by using acetate and propionate as carbon sources. Appl Biochem Biotechnol. 121:289–301. doi:10.1385/ABAB:121:13:0289.

Iwata T. 2015. Biodegradable and biobased polymers: future prospects of ecofriendly plastics. Angew Chem Int Ed. 54(11):3210–3215. doi:10.1002/anie.201410770.

Jeffrey GA. 1997. An introduction to hydrogen bonding, volume 12. New York: Oxford university press. Kihara T, Hiroe A, Ishii HM, Mizuno K, Tsuge T. 2017. Bacillus cereus type polyhydroxyalkanoate biosynthetic gene cluster contains Rspecific enoylCoA hydratase gene. Biosci Biotechnol Biochem. 81(8):1627–1635. doi:10.1080/09168451.2017.1325314.

Kim J, Kim YJ, Choi SY, Lee SY, J KK. 2017. Crystal structure of Ralstonia eutropha polyhydroxyalkanoate synthase C terminal domain and reaction mechanisms. Biotechnol J. 12(1):1600648. doi:10.1002/biot.201600648.

Kumar S, Stecher G, Tamura K. 2016. MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol. 33(7):1870–1874. doi:10.1093/molbev/msw054.

Laemmli UK. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 227(5259):680–685. doi:10.1038/227680a0.

Liebergesell M, Steinbuchel A. 1992. Cloning and nucleotide sequences of genes relevant for biosynthesis of poly(3hydroxybutyric acid) in Chromatium vinosum strain D. Eur J Biochem. 209(1):135–150. doi:10.1111/j.14321033.1992.tb17270.x.

Liu Z, Zhu Z, Yang J, Wu S, Liu Q, Wang M, Cheng H, Yan J, L W. 2019. Domaincentric dissection and classification of prokaryotic poly (3hydroxyalkanoate) synthases. bioRxiv p. 693432. doi:10.1101/693432.

Mesquita DP, Amaral AL, Leal C, Oehmen A, Reis MAM, Ferreira EC. 2015. Polyhydroxyalkanoate granules quantification in mixed microbial cultures using image analysis: Sudan Black B versus Nile Blue A staining. Anal Chim Acta. 865:8–15. doi:10.1016/j.aca.2015.01.018.

Mezolla V, D’Urso OF, Poltronieri P. 2018. Role of PhaC type I and type II enzymes during PHA biosynthesis. Polymers. 10(910):1–12. doi:10.3390/polym10080910.

Morris AL, MacArthur MW, Hutchinson EG, Thornton JM. 1992. Stereochemical quality of protein structure coordinates. Proteins: Struct Funct Bioinf. 12(4):345–364. doi:10.1002/prot.340120407.

Müh U, Sinskey AJ, Kirby DP, S LW, Stubbe JA. 1999. PHA synthase from Chromatium vinosum: cysteine 149 is involved in covalent catalysis. Biochemistry. 38(2):826–837. doi:10.1021/bi9818319.

Ramsay BA, Lomaliza K, Chavarie C, Dube B, Bataille P, Ramsay JA. 1990. Production of poly(betahydroxybutyriccobetahydroxyvaleric) acids. Appl Environ Microbiol. 56(7):2093–2098. doi:10.1128/AEM.56.7.20932098.

Roussel A, Miled N, BertiDupuis L, Rivière M, Spinelli S, Berna P, Gruber V, Verger R, Cambillau C. 2002. Crystal structure of the open form of dog gastric lipase in complex with a phosphonate inhibitor. J Biol Chem. 277(3):2266–2274. doi:10.1074/jbc.M109484200.

Sagong HY, Son HF, Choi SY, Lee SY, J KK. 2018. Structural insight into polyhydroxyalkanoates biosynthesis. Trends Biochem Sci. 43(10):790–805. doi:10.1016/j.tibs.2018.08.005.

Saitou N, Nei M. 1987. The neighborjoining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol. 4(4):406–425. doi:10.1093/oxfordjournals.molbev.a040454.

Sudesh K, Abe H, Doi Y. 2000. Synthesis, structure and properties of polyhydroxyalkanoates: biological polyesters. Prog Polym Sci. 25(10):1503–1555. doi:10.1016/S00796700(00)000356.

Tian J, Sinskey AJ, Stubbe JA. 2005. Detection of intermediates from the polymerization reaction catalyzed by a D302A mutant of class III polyhydroxyalkanoate (PHA) synthase. Biochem. 44(5):1495–1503. doi:10.1021/bi047734z.

Tsuge T, Hyakutake M, Mizuno K. 2015. Class IV polyhydroxyalkanoate (PHA) synthases and PHAproducing Bacillus. Appl Microbiol Biotechnol. 99(15):6231– 6240. doi:10.1007/s0025301567779.

Webb B, Sali A. 2016. Comparative protein structure modeling using MODELLER. Curr Protoc Bioinf.54(1):5–6. doi:10.1002/cpbi.3.

Wittenborn EC, Jost M, Wei Y, A SJ, Drennan CL. 2016. Structure of the catalytic domain of the class I polyhydroxybutyrate synthase from Cupriavidus necator. J Biol Chem. 291(48):25264–25277. doi:10.1074/jbc.M116.756833.

Yang J, Yan R, Roy A, Xu D, J P, Zhang Y. 2015. The ITASSER Suite: protein structure and function prediction. Nat. Methods 12(1):7. doi:10.1038/nmeth.3213.

Yang J, Zhang Y. 2015. ITASSER server: new development for protein structure and function predictions. Nucleic Acids Res. 43(W1):W174–W181. doi:doi.org/10.1093/nar/gkv342.

Yanti NA, Sembiring L, S M. 2009. Production of Polyα hydroxybutyrate (PHB) from Sago Starch by The Native Isolate Bacillus megaterium PSA10. IJ Biotech. 11(1):1111–1116. doi:10.22146/ijbiotech.7804.

Zhang X, Luo R, Wang Z, Deng Y, Q CG. 2009. Application of (R)3hydroxyalkanoate methyl esters derived from microbial polyhydroxyalkanoates as novel biofuels. Biomacromol. 10(4):707–711. doi:10.1021/bm801424e.

Zuckerkandl E, Pauling L. 1965. Evolutionary divergence and convergence in proteins. In: V Bryson, HJ Vogel, editors, Evolving genes and proteins. Amsterdam: Elsevier. p. 97–166. doi:10.1016/B978148322734 4.500176.

DOI: https://doi.org/10.22146/ijbiotech.53717