Cloning, Expression, and Characterization of a Cold-Adapted Shikimate Kinase from the Psychrophilic Bacterium Colwellia psychrerythraea 34H
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Most cold-adapted enzymes possess higher Km and kcat values than those of their mesophilic counterparts to maximize the reaction rate. This characteristic is often ascribed to a high structural flexibility and improved dynamics in the active site. However, this may be less convincing to cold-adapted metabolic enzymes, which work at substrate concentrations near Km. In this respect, cold adaptation of a shikimate kinase (SK) in the shikimate pathway from psychrophilic Colwellia psychrerythraea (CpSK) was characterized by comparing it with a mesophilic Escherichia coli homolog (EcSK). The optimum temperatures for CpSK and EcSK activity were approximately 30°C and 40°C, respectively. The melting points were 33°C and 45°C for CpSK and EcSK, respectively. The ΔGH2O (denaturation in the absence of denaturing agent) values were 3.94 and 5.74 kcal/mol for CpSK and EcSK, respectively. These results indicated that CpSK was a cold-adapted enzyme. However, contrary to typical kinetic data, CpSK had a lower Km for its substrate shikimate than most mesophilic SKs, and the kcat was not increased. This observation suggested that CpSK may have evolved to exhibit increased substrate affinity at low intracellular concentrations of shikimate in the cold environment. Sequence analysis and homology modeling also showed that some important salt bridges were lost in CpSK, and higher Arg residues around critical Arg 140 seemed to increase flexibility for catalysis. Taken together, these data demonstrate that CpSK exhibits characteristics of cold adaptation with unusual kinetic parameters, which may provide important insights into the cold adaptation of metabolic enzymes.
Abstract
Most cold-adapted enzymes possess higher Km and kcat values than those of their mesophilic counterparts to maximize the reaction rate. This characteristic is often ascribed to a high structural flexibility and improved dynamics in the active site. However, this may be less convincing to cold-adapted metabolic enzymes, which work at substrate concentrations near Km. In this respect, cold adaptation of a shikimate kinase (SK) in the shikimate pathway from psychrophilic Colwellia psychrerythraea (CpSK) was characterized by comparing it with a mesophilic Escherichia coli homolog (EcSK). The optimum temperatures for CpSK and EcSK activity were approximately 30°C and 40°C, respectively. The melting points were 33°C and 45°C for CpSK and EcSK, respectively. The ΔGH2O (denaturation in the absence of denaturing agent) values were 3.94 and 5.74 kcal/mol for CpSK and EcSK, respectively. These results indicated that CpSK was a cold-adapted enzyme. However, contrary to typical kinetic data, CpSK had a lower Km for its substrate shikimate than most mesophilic SKs, and the kcat was not increased. This observation suggested that CpSK may have evolved to exhibit increased substrate affinity at low intracellular concentrations of shikimate in the cold environment. Sequence analysis and homology modeling also showed that some important salt bridges were lost in CpSK, and higher Arg residues around critical Arg 140 seemed to increase flexibility for catalysis. Taken together, these data demonstrate that CpSK exhibits characteristics of cold adaptation with unusual kinetic parameters, which may provide important insights into the cold adaptation of metabolic enzymes.
참고문헌 (59)
- Ásgeirsson B, "Microscopic rate-constants for substrate binding and acylation in cold-adaptation of trypsin I from Atlantic cod", FEBS Lett, 580, pp.4639~4644 (2006)
- Bentahir M, "Structural, kinetic, and calorimetric characterization of the cold-active phosphoglycerate kinase from the antarctic Pseudomonas sp", TACII18. J. Biol. Chem, 275, pp.11147~11153 (2000)
- Cerasoli E, "The refolding of type II shikimate kinase from Erwinia chrysanthemi aft er denat urat ion in urea", Eur. J. Biochem, 269, pp.2124~2132 (2002)
- Cheng W-C, "Structural basis for shikimate-binding specificity of Helicobacter pylori shikimate kinase", J. Bacteriol, pp.8156~8163 (2005)
- Cheng W-C, "Structures of Helicobacter pylori shikimate kinase reveal a selective inhibitor-induced-fit mechanism", PLoS One, 7, pp.e33481~ (2012)
- Collins T, "A novel family 8 xylanase, functional and physicochemical characterization", J. Biol. Chem, 277, pp.35133~35139 (2002)
- Coracini JD, "Shikimate kinase, a protein target for drug design", Curr. Med. Chem, 21, pp.592~604 (2014)
- D’Amico S, "Molecular basis of cold adaptation", Philos. Trans. R. Soc. Lond. B Biol. Sci, 357, pp.917~925 (2002)
- D’Amico S, "Psychrophilic microorganisms: challenges for life", EMBO Rep, 7, pp.385~389 (2006)
- DeFeyt er RC, "Purificat ion and propert ies of shikimate kinase II from Escherichia coli K-12", J. Bacteriol, 165, pp.331~333 (1986)
- "The PyMOL Molecular Graphics System" (2002)[URL]
- De Santi C, "A new alkaliphilic cold-active esterase from the psychrophilic marine bacterium Rhodococcus sp.: functional and structural studies and biotechnological potential", Appl. Biochem. Biotechnol, 172, pp.3054~3068 (2014)
- Di Tommaso P, "T-Coffee: a Web server for the multiple sequence alignment of protein and RNA sequences using structural information and homology extension", Nucleic Acids Res, 39, pp.W13~W17 (2011)
- Feller G, "Psychrophilic enzymes: molecular basis of cold adaptation", Cell. Mol. Life Sci, 53, pp.830~841 (1997)
- Feller G, "Enzymes from psychrophilic organisms", FEMS Microbiol. Rev, 18, pp.189~202 (1996)
- Feller G, "Enzymes from cold-adapted microorganisms - t he class C beta-lactamase from the antarctic psychrophile Psychrobacter Immobilis A5", Eur. J. Biochem, 244, pp.186~191 (1997)
- Fucile G, "Structural and biochemical investigation of two Arabidopsis shikimat e kinases: t he heat -inducible isoform is thermostable", Protein Sci, 20, pp.1125~1136 (2011)
- Gan J, "Cryst al st ruct ure of Mycobacterium tuberculosis shikimate kinase in complex with shikimic acid and an ATP analogue", Biochemistry, 45, pp.8539~8545 (2006)
- Genicot S, "Trypsin from antarctic fish (Paranotothenia magellanica forster) as compared with trout (Salmo gairdneri) trypsin", Comp. Biochem. Physiol. B, 90, pp.601~609 (1988)
- Georlette D, "Physics and Chemistry Basis of Biotechnology" (2002), Springer Netherlands, Dordrecht
- Greenfield NJ, "Using circular dichroism collected as a function of temperature to determine the thermodynamics of protein unfolding and binding interactions", Nat. Protoc, 1, pp.2527~2535 (2006)
- Han C, "Discovery of Helicobacter pylori shikimate kinase inhibitors:bioassay and molecular modeling", Bioorg. Med. Chem, 15, pp.656~662 (2007)
- Herrmann KM, "The shikimate pathway", Annu. Rev. Plant Physiol. Plant Mol. Biol, 50, pp.473~503 (1999)
- Hoyoux A, "Cold-adapted beta-galactosidase from the Antarctic psychrophile Pseudoalteromonas haloplanktis", Appl. Environ. Microbiol, 67, pp.1529~1535 (2001)
- Jahandideh M, "Elucidating the protein cold-adaptation: investigation of the parameters enhancing protein psychrophilicity", J. Theor. Biol, 255, pp.113~118 (2008)
- Kaufmman S, "Methods in Enzymology, Vol. 147:Metabolism of Aromatic Amino Acids and Amines" (1987), Academic Press
- Kim S-Y, "Structural basis for cold adaptation: sequence, biochemical properties, and crystal structure of malate dehydrogenase from a psychrophile Aquaspirillium arcticum", J. Biol. Chem, 274, pp.11761~11767 (1999)
- 김영옥, "A Novel Esterase from Paenibacillus sp. PBS-2 Is a New Member of the β-Lactamase Belonging to the Family VIII Lipases", Esterases, 2014, pp.24(9), 1260~1268[DOI]
- Krell T, "Biochemical and X-ray crystallographic studies on shikimate kinase: the important structural role of the P-loop lysine", Protein Sci, 10, pp.1137~1149 (2001)
- Kulakova L, "Cold-active esterase from Psychrobacter sp. Ant 300:gene cloning, characterization, and the effects of Gly → Pro substitution near the active site on its catalytic activity and stability", Biochim. Biophys. Acta, 1696, pp.59~65 (2004)
- Kumar M, "COPid: composition based protein identification", In Silico Biol, 8, pp.121~128 (2008)
- Lakowicz JR, "Principles of Fluorescence Spectroscopy" (2006), Springer US
- 이용석, "A Cold-Adapted Carbohydrate Esterase from the Oil-Degrading Marine Bacterium Microbulbifer thermotolerans DAU221: Gene Cloning, Purification, and Characterization", Journal of Microbiology and Biotechnology, 24(7), pp.925~935 (2014), 한국미생물·생명공학회[DOI]
- Leiros I, "The structure of uracil-DNA glycosylase from Atlantic cod (Gadus morhua) reveals cold-adaptation features", Acta Crystallogr. D Biol. Crystallogr, 59, pp.1357~1365 (2003)
- Shangyong Li, "Cloning, Expression, and Characterization of a Cold-Adapted and Surfactant-Stable Alginate Lyase from Marine Bacterium Agarivorans sp. L11", Journal of Microbiology and Biotechnology, 25(5), pp.681~686 (2015), 한국미생물·생명공학회[DOI]
- Maiangwa J, "Adaptational properties and applications of cold-active lipases from psychrophilic bacteria", Extremophiles, 19, pp.235~247 (2015)
- Metpally RPR, "Comparative proteome analysis of psychrophilic versus mesophilic bacterial species : insights into the molecular basis of cold adaptation of proteins", BMC Genomics, 10, pp.11~ (2009)
- Moyer CL, "Psychrophiles and psychrotrophs", ~ (2007)
- Pace CN, "Protein structure, stability and solubility in water and other solvents", Philos. Trans. R. Soc. Lond. B Biol. Sci, 359, pp.1225~1234 (2004)
- Paredes DI, "Comparative void-volume analysis of psychrophilic and mesophilic enzymes: structural bioinformatics of psychrophilic enzymes reveals sources of core flexibility", BMC Struct. Biol, 11, pp.42~ (2011)
- Robert X, "Deciphering key features in protein structures with the new ENDscript server", Nucleic Acids Res, 42, pp.W320~W324 (2014)
- Romanowski MJ, "Crystal structure of the Escherichia coli shikimate kinase I(AroK)that confers sensitivity to mecillinam", Proteins, 47, pp.558~562 (2002)
- Rosado LA, "The mode of action of recombinant Mycobacterium tuberculosis shikimate kinase:kinetics and thermodynamics analyses", PLoS One, 8, pp.e61918~ (2013)
- Siddiqui KS, "Cold-adapted enzymes", Annu. Rev. Biochem, 75, pp.403~433 (2006)
- Siddiqui KS, "Role of lysine versus arginine inenzyme cold-adaptation : modifying lysine to homo-arginine stabilizes the cold-adapted α-amylase from Pseudoalteramonas haloplanktis", Proteins, 64, pp.486~501 (2006)
- Simithy J, "Development of an ESI-LC-MS-based assay for kinetic evaluation of Mycobacterium tuberculosis shikimate kinase activity and inhibition", Anal. Chem, 87, pp.2129~2136 (2015)
- Simithy J, "Identification of shikimate kinase inhibitors among anti-Mycobacterium tuberculosis compounds by LC-MS", Tuberculosis (Edinb.), 94, pp.152~158 (2014)
- Simpson BK, "Purification and characterization of trypsin from the Greenland cod (Gadus ogac). 1. Kinetic and thermodynamic characteristics", Can. J. Biochem. Cell Biol, 62, pp.894~900 (2011)
- Singh P, "Diversity, cold active enzymes and adaptation strategies of bacteria inhabiting glacier cryoconite holes of High Arctic", Extremophiles, 18, pp.229~242 (2014)
- Smalås AO, "Cold adapted enzymes", Biotechnol. Annu. Rev, 6, pp.1~57 (2000)
- Struvay C, "Optimization to low temperature activity in psychrophilic enzymes", Int. J. Mol. Sci, 13, pp.11643~11665 (2012)
- Hongfei Su, "Cloning, Expression, and Characterization of a Cold-Active and Organic Solvent-Tolerant Lipase from Aeromicrobium sp. SCSIO 25071", Journal of Microbiology and Biotechnology, 26(6), pp.1067~1076 (2016), 한국미생물·생명공학회[DOI]
- Sutton KA, "Structure of shikimate kinase, an in vivo essential metabolic enzyme in the nosocomial pathogen Acinetobacter baumannii, in complex with shikimate", Acta Crystallogr. D Biol. Crystallogr, 71, pp.1736~1744 (2015)
- Szilágyi A, "Structural differences between mesophilic, moderately thermophilic and extremely thermophilic protein subunits : results of a comprehensive survey", Structure, 8, pp.493~504 (2000)
- Tang MAK, "Cold adaptation:structural and functional characterizations of psychrophilic and mesophilic acetate kinase", Protein J, 33, pp.313~322 (2014)
- Wang G, "A novel cold-adapted and highly salt-tolerant esterase from Alkalibacterium sp. SL3 from t he sediment of a soda lake", Sci. Rep, 6, pp.19494~ (2016)
- Webb B, "Comparative protein structure modeling using MODELLER", Curr. Protoc. Bioinformatics, 47, pp.5.6.1~5.6.32 (2014)
- Xu Y, "Metabolic enzymes from psychrophilic bacteria: challenge of adaptation to low temperatures in ornithine carbamoyltransferase from Moritella abyssi", J. Bacteriol, 185, pp.2161~2168 (2003)
- Zanphorlin LM, "Oligomerization as a strategy for cold adaptation: structure and dynamics of the GH1 β-glucosidase from Exiguobacterium antarcticum B7", Sci. Rep, 6, pp.23776~ (2016)