Characteristics and Function of the Chitin Binding Protein from Xenorhabdus nematophila

Author(s): Jia Liu, Ping Song, Jie Zhang, Ziyan Nangong, Xiaobei Liu, Yue Gao, Qinying Wang*.

Journal Name: Protein & Peptide Letters

Volume 26 , Issue 6 , 2019

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Abstract:

Background: Genome sequence analysis (GenBank access No.: FN667742.1) shows that Xenorhabdus nematophila ATCC19061 contains one gene (Xn-cbp) encoding chitin binding protein (Xn-CBP).

Objective: The present work aims to clarify the characteristics and function of Xn-CBP from X. nematophila HB310.

Methods: In this study, the Xn-cbp gene was cloned and expressed in Escherichia coli BL21 (DE3). Substrate binding assays were performed to explain the ability of Xn-CBP combined with the polysaccharide. The insecticidal toxicity of Xn-CBP against the second-instar larvae of Helicoverpa armigera was determined by feeding method. Besides, the antifungal activity of Xn-CBP against Coniothyrium diplodiella, Verticillium dahlia, and Fusarium oxysporum was tested by spore germination assay and hyphal extension assay.

Results: Xn-CBP encoded 199 amino acids with a calculated mass of 28 kDa, which contained a signal peptide and a chitin binding domain. The Bmax and Kd values of Xn-CBP to colloidal chitin were 2.46 and 4.08, respectively. Xn-CBP had insecticidal activity against the H. armigera with a growth inhibition rate of 84.08%. Xn-CBP had the highest spore germination inhibitory effect on C. diplodiella with the inhibition rate of 83.11%. The hyphal growth inhibition rate of Xn-CBP to F. oxysporum, 41.52%, was higher than the other two fungi.

Conclusion: The Xn-CBP had the highest binding ability to colloidal chitin and it showed insecticidal activity and antifungal activity. The present study laid a foundation for further exploitation and utilization of X. nematophila.

Keywords: Antifungal activity, chitin binding protein, insecticidal activity, substrate binding, Xenorhabdus nematophila, spore germination.

[1]
Bhuvanachandra, B.; Madhuprakash, J.; Podile, A.R. Active-site mutations improved the transglycosylation activity of Stenotrophomonas maltophilia chitinase A. BBA.-. Proteins Proteom., 2018, 1866, 407-414.
[2]
Gao, L.; Sun, J.; Secundo, F.; Gao, X.; Xue, C.; Mao, X. Cloning, characterization and substrate degradation mode of a novel chitinase from Streptomyces albolongus ATCC 27414. Food Chem., 2018, 261, 329-336.
[3]
Shinoda, T.; Kobayashi, J.; Matsui, M.; Chinzei, Y. Cloning and functional expression of a chitinase cDNA from the common cutworm, Spodoptera litura, using a recombinant baculovirus lacking the virus-encoded chitinase gene. Insect Biochem. Mol. Biol., 2001, 31, 521-532.
[4]
Frederiksen, R.F.; Paspaliari, D.K.; Larsen, T.; Storgaard, B.G.; Larsen, M.H.; Ingmer, H.; Palcic, M.M.; Leisner, J.J. Bacterial chitinases and chitin-binding proteins as virulence factors. Microbiology, 2013, 159(Pt5), 833-847.
[5]
Ikeda, M.; Kondo, Y.; Matsumiya, M. Purification, characterization, and molecular cloning of chitinases from the stomach of the threeline grunt Parapristipoma trilineatum. Process Biochem., 2013, 48, 1324-1334.
[6]
Shehata, A.N.; Abd El Aty, A.A.; Darwish, D.A.; Wahab, W.A.A.; Mostafa, F.A. Purification, physicochemical and thermodynamic studies of antifungal chitinase with production of bioactive chitosan-oligosaccharide from newly isolated Aspergillus griseoaurantiacus KX010988. Int. J. Biol. Macromol., 2018, 107, 990-999.
[7]
Mehmood, M.A.; Latif, M.; Hussain, K.; Gull, M.; Latif, F.; Rajoka, M.I. Heterologous expression of the antifungal beta-chitin binding protein CBP24 from Bacillus thuringiensis and its synergistic action with bacterial chitinases. Protein Pept. Lett., 2015, 22, 39-44.
[8]
Merino, S.T.; Cherry, J. Progress and challenges in enzyme development for biomass utilization. Adv. Biochem. Eng. Biotechnol., 2007, 108, 95-120.
[9]
Purushotham, P.; Arun, P.V.P.S.; Prakash, J.S.S.; Podile, A.R. Chitin binding proteins act synergistically with chitinases in Serratia proteamaculans 568. PLoS One, 2012, 7e367145
[10]
Yin, J.; Yang, S.; Li, K.; Guo, W.; Cao, Y. Identification and molecular characterization of a chitin-binding protein from the Beet Webworm, Loxostege sticticalis L. Int. J. Mol. Sci., 2014, 15, 19147-19161.
[11]
Gifoni, J.M.; Oliveira, J.T.A.; Oliveira, H.D.; Batista, A.B.; Pereira, M.L.; Gomes, A.S.; Oliveira, H.P.; Grangeiro, T.B.; Vasconcelos, I.M. A novel chitin-binding protein from Moringa oleifera seed with potential for plant disease control. Biopolymers, 2012, 98, 406-415.
[12]
Mehmood, M.A.; Xiao, X.; Hafeez, F.Y.; Gai, Y.; Wang, F. Molecular characterization of the modular chitin binding protein Cbp50 from Bacillus thuringiensis serovar konkukian. Antonie van Leeuwenhoek, 2011, 100, 445-453.
[13]
Ingrid Gutierrez-Roman, M.; Dunn, M.F.; Tinoco-Valencia, R.; Holguin-Melendez, F.; Huerta-Palacios, G.; Guillen-Navarro, K. Potentiation of the synergistic activities of chitinases ChiA, ChiB and ChiC from Serratia marcescens CFFSUR-B2 by chitobiase (Chb) and Chitin Binding Protein (CBP). World J. Microb. Biot., 2014, 30, 33-42.
[14]
Manjeet, K.; Purushotham, P.; Neeraja, C.; Podile, A.R. Bacterial chitin binding proteins show differential substrate binding and synergy with chitinases. Microbiol. Res., 2013, 168, 461-468.
[15]
Vaaje-Kolstad, G.; Houston, D.R.; Riemen, A.; Eijsink, V.; van Aalten, D. Crystal structure and binding properties of the Serratia marcescens chitin-binding protein CBP21. J. Biol. Chem., 2005, 280, 11313-11319.
[16]
Mehmood, M.A.; Xiao, X.; Hafeez, F.Y.; Gai, Y.; Wang, F. Molecular characterization of an endochitinase from Bacillus thuringiensis subsp konkukian. World J. Microb. Biot., 2010, 26, 2171-2178.
[17]
Mehmood, M.A.; Latif, M.; Hafeez, F.Y. Heterologous expression and characterization of an antifungal chitinase Chi39 from Bacillus thuringiensis serovar konkukian. Pakistan. J. Life Soc. Sci., 2012, 10, 116-122.
[18]
Mehmood, M.A.; Hussain, K.; Latif, F.; Tabassum, M.R.; Gull, M.; Gill, S.S.; Saqib, A.; Iqbal, Z. Synergistic action of the antifungal beta-chitin binding protein CBP50 from Bacillus thuringiensis with bacterial chitinases. Curr. Proteomics, 2014, 11, 23-26.
[19]
Joshi, M.C.; Sharma, A.; Kant, S.; Birah, A.; Gupta, G.P.; Khan, S.R.; Bhatnagar, R.; Banerjee, N. An insecticidal GroEL protein with chitin binding activity from Xenorhabdus nematophila. J. Biol. Chem., 2008, 283, 28287-28296.
[20]
Yang, Q.; Zhang, J.; Li, T.; Liu, S.; Song, P.; Nangong, Z.; Wang, Q. PirAB protein from Xenorhabdus nematophila HB310 exhibits a binary toxin with insecticidal activity and cytotoxicity in Galleria mellonella. J. Invertebr. Pathol., 2017, 148, 43-50.
[21]
Chen, G.; Zhang, Y.; Li, J.; Dunphy, G.B.; Punja, Z.K.; Webster, J.M. Chitinase activity of Xenorhabdus and Photorhabdus species, bacterial associates of entomopathogenic nematodes. J. Invertebr. Pathol., 1996, 68, 101-108.
[22]
Ffrench-Constant, R.; Bowen, D. Photorhabdus toxins: Novel biological insecticides. Curr opin microbiol., 1999, 2, 284-288.
[23]
Wang, Y.H.; Feng, J.T.; Zhang, Q.; Zhang, X. Optimization of fermentation condition for antibiotic production by Xenorhabdus nematophila with response surface methodology. J. Appl. Microbiol., 2008, 104, 735-744.
[24]
Wang, Q.; Nangong, Z.; Yang, J.; Song, P.; Wang, Y.; Cui, L.; Cui, L. Toxic activity of a protein complex purified from Xenorhabdus nematophila HB310 to Plutella xylostella larvae. Insect Sci., 2012, 19, 329-336.
[25]
Bernard, D.; Quatannens, B.; Vandenbunder, B.; Abbadie, C. Rel/NF-kappa B transcription factors protect against Tumor Necrosis Factor (TNF)-related apoptosis-inducing ligand (TRAIL)-induced apoptosis by up-regulating the TRAIL decoy receptor DcR1. J. Biol. Chem., 2001, 276, 27322-27328.
[26]
Kirubakaran, S.I.; Sakthivel, N. Cloning and overexpression of antifungal barley chitinase gene in Escherichia coli. Protein Expres. Purif., 2007, 52, 159-166.
[27]
Yang, J.; Zeng, H.; Lin, H.; Yang, X.; Liu, Z.; Guo, L.; Yuan, J.; Qiu, D. An insecticidal protein from Xenorhabdus budapestensis that results in prophenoloxidase activation in the wax moth, Galleria mellonella. J. Invertebr. Pathol., 2012, 110, 60-67.
[28]
Saito, A.; Miyashita, K.; Biukovic, G.; Schrempf, H. Characteristics of a Streptomyces coelicolor A3(2) extracellular protein targeting chitin and chitosan. Appl. Environ. Microb., 2001, 67, 1268-1273.
[29]
Vaaje-Kolstad, G.; Bohle, L.A.; Gaseidnes, S.; Dalhus, B.; Bjoras, M.; Mathiesen, G.; Eijsink, V.G.H. Characterization of the chitinolytic machinery of Enterococcus faecalis V583 and high-resolution structure of its Oxidative CBM33 Enzyme. J. Mol. Biol., 2012, 416, 239-254.
[30]
Chandrasekaran, R.; Revathi, K.; Thanigaivel, A.; Kirubakaran, S.A.; Senthil-Nathan, S. Bacillus subtilis chitinase identified by matrix-assisted laser desorption/ionization time-of flight/time of flight mass spectrometry has insecticidal activity against Spodoptera litura Fab. Pestic. Biochem. Phys., 2014, 116, 1-12.
[31]
Suganthi, M.; Senthilkumar, P.; Arvinth, S.; Chandrashekara, K.N. Chitinase from Pseudomonas fluorescens and its insecticidal activity against Helopeltis theivora. J. Gen. Appl. Microbiol., 2017, 63, 222-227.
[32]
Downing, K.J.; Leslie, G.; Thomson, J.A. Biocontrol of the sugarcane borer Eldana saccharina by expression of the Bacillus thuringiensis cry1Ac7 and Serratia marcescens chiA genes in sugarcane-associated bacteria. Appl. Environ. Microb., 2000, 66, 2804-2810.
[33]
Eleazar Barboza-Corona, J.; Luis Delgadillo-Angeles, J.; Cristobal Castaneda-Ramirez, J.; Eleazar Barboza-Perez, U.; Edith Casados-Vazquez, L.; Bideshi, D.K.; Cristina Del Rincon-Castro, M. Bacillus thuringiensis subsp kurstaki HD1 as a factory to synthesize alkali-labile ChiA74 [increment] sp chitinase inclusions, Cry crystals and spores for applied use. Microb. Cell Fact., 2014, 13, 15.
[34]
Thamthiankul, S.; Moar, W.J.; Miller, M.E.; Panbangred, W. Improving the insecticidal activity of Bacillus thuringiensis subsp aizawai against Spodoptera exigua by chromosomal expression of a chitinase gene. Appl. Microbiol. Biot., 2004, 65, 183-192.
[35]
Ajit, N.S.; Verma, R.; Shanmugam, V. Extracellular chitinases of Fluorescent pseudomonads antifungal to Fusarium oxysporum f. Sp dianthi causing carnation wilt. Curr. Microbiol., 2006, 52, 310-316.
[36]
Lu, Y.; Wang, N.; He, J.; Li, Y.; Gao, X.; Huang, L.; Yan, X. Expression and characterization of a novel chitinase with antifungal activity from a rare actinomycete, Saccharothrix yanglingensis Hhs.015. Protein Expres. Purif., 2018, 143, 45-51.
[37]
Mander, P.; Cho, S.S.; Choi, Y.H.; Panthi, S.; Choi, Y.S.; Kim, H.M.; Yoo, J.C. Purification and characterization of chitinase showing antifungal and biodegradation properties obtained from Streptomyces anulatus CS242. Arch. Pharm. Res., 2016, 39, 878-886.
[38]
Arora, N.; Sachdev, B.; Gupta, R.; Vimala, Y.; Bhatnagar, R.K. Characterization of a chitin-binding protein from Bacillus thuringiensis HD-1. PLoS One, 2013, 8e666036


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Article Details

VOLUME: 26
ISSUE: 6
Year: 2019
Page: [414 - 422]
Pages: 9
DOI: 10.2174/0929866526666190327143335
Price: $65

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