Bazı Kimyasalların Etkisi Altında Escherichia coli’nin OmpF - OmpC Porin Protein Sentezi ve Bu Sentezde EnvZ, RpoS, H-NS, AcP’ın Rolünün Belirlenmesi

Yıl 2018, Cilt: 11 Sayı: 2, 285 - 299, 31.08.2018

Cihan Darcan , Hülya Yılmaz

https://doi.org/10.18185/erzifbed.376759

Öz

Escherichia coli’nin dış membranında
bulunan OmpC ve OmpF porin proteinleri, stres koşullarına karşı korunmasında
önemli yer tutar. Bu çalışmada dezenfektan özellikli çeşitli kimyasal
maddelerin E. coli’nin OmpC ve OmpF
porin protein sentezini nasıl etkilediği araştırılmıştır. E. coli’de, formaldehit, klor,
etanol, çamaşır suyu, H₂O₂ ve Sodyum Dodesil Sülfatın
(SDS) belirlenen konsatrasyonları besiyerine eklenerek, ompC ve ompF genleri yerine raportör gen olarak eklenen β-galaktosidaz’ın aktivitesi spektrofotometrik olarak ölçülmüş ve elde
edilen sonuçlar kontrol grupları ile karşılaştırılmıştır. Sonuç olarak
çalışılan kimyasal maddelerin E. coli’nin
büyümesini oldukça yavaşlattığı, ancak yinede üremenin gerçekleştiği
belirlenmiştir. E. coli’de OmpF
sentezinin formaldehit, etanol ve SDS ilave edildikten 2 saat sonra oldukça
azaldığı, çamaşır suyu ve klor ilavesinde biraz azaldığı ve H₂O₂
ilave edildiği zaman ise değişmediği tespit edilmiştir. OmpC sentezinde
ise klor, formaldehit, H₂O₂ ilave edildiğinde %50
oranında azaldığı belirlenirken, çamaşır suyunda yaklaşık 2 kat ve SDS ilave
edilen örneklerde ise 1.5 katlık bir artış belirlenmiştir. Ayrıca hem OmpC hem
de OmpF porin proteininin sentezinde RpoS, Pta, EnvZ ve H-NS proteinlerinin
rollerinin olduğu belirlenmiştir. Ancak moleküler mekanizmaları henüz
aydınlatılamamıştır.

Anahtar Kelimeler

Porin, Dezenfektan kimyasallar, Escherichia coli, Porin

The Role of EnvZ, RpoS, H-NS, AcP and Synthesis of OmpC-OmpF Porin Proteins of Escherichia coli Under Effect of Some Chemicals

Öz

OmpC and OmpF porin proteins, located on the
outer membrane of Escherichia coli,
play an important role in protection against stress conditions. In this study, it was investigated how various chemical substances as a
disinfectant affect E. coli OmpC and
OmpF porin protein synthesis. The activity of β-galactosidase, added as a
reporter gene in place of ompC and ompF genes, was measured
spectrophotometrically by adding formaldehyde, chlorine, ethanol, bleach, H₂O₂
and Sodium Dodecyl Sulfate (SDS), used as disinfectants in E. coli. The results were compared with control groups. As a result, it was determined that the studied chemical substances
slowed down the growth of E. coli at
the added concentrations, but continued to multiply. It was determined that the
OmpF synthesis in E. coli decreased
considerably after 2 hours from the addition of formaldehyde, ethanol and SDS,
decreased slightly during the addition of bleach and chlorine, and did not
change when H₂O₂ was added. In the OmpC synthesis, when
chlorine, formaldehyde, and H₂O₂ were added, it was
determined to decrease by 50% compared to the untreated state, while the
increase was found as a 1.5 time in the cases where SDS was added and about 2
times in the bleach. In these alterations, it has been determined that the
roles of RpoS, Pta, EnvZ, and H-NS proteins on both OmpC and OmpF porin protein
synthesis but have not yet been elucidated by their molecular mechanisms.

Anahtar Kelimeler

Disinfectan chemicals, Escherichia coli, Porin

Kaynakça

• Achouak, W., Pages, J. M., De Mot, R., Molle, G. and Heulin T., 1998. A major outer membrane protein of Rahnella aquatilis functions as a porin and root adhesin. Journal of Bacteriology, 180, 900-913.
• Achouak, W., Heulin, T. and Pages, J. M., 2001. Multiple facets of bacterial porins. FEMS Microbiology Letters, 199, 1-7.
• Aguayo, D., Pacheco, N., Morales, E.H., Collao, B., Luraschi, R., Cabezas, C., Calderón, P., González-Nilo, F., Gil, F., Calderón, I.L. et .al 2015. Hydrogen peroxide and hypochlorous acid influx through the major S. typhimurium porin OmpD is affected by substitution of key residues of the channel. Archives of Biochemistry and Biophysics, 568, 38-45.
• Aiba, H., Matsuyama, S. I., Mizuno, T. ve Mizushima, S., 1987. Function of micF as an antisense RNA in osmoregulatory expression of the ompF gene in E. coli. Journal of Bacteriology, 169 (7), 3007-3012.
• Alekshun, M.N. ve Levy, S.B., 1999. The mar regulon: multiple resistance to antibiotics and other toxic chemicals. Trends in Microbiology, 7 (10), 410-413.
• Andersen, J., Forst, S. A., Zhao, K., Inouye, M. ve Delihas, N., 1989. The function of micF RNA: micF RNA is a major factor in the thermal regulation of OmpF protein in Escherichia coli. The Journal of Biological Chemistry, 264, 17961-17970.
• Barrette, W.C., Hannum, D.M., Wheeler, W.D. ve Hurst, J.K., 1989. General mechanism for the bacterial toxicity of hypochlorous acid: abolition of ATP production. Biochemistry, 28, 9172–9178.
• Basle, A., Qutub, R., Mehrazin, M., Wibbenmeyer, J. ve Delcour, A.H., 2004. Deletions of single extracellular loops affect pH sensitivity, but not voltage dependence, of the Escherichia coli porin OmpF. Protein Engineering Design & Selection, 17, 665–672.
• Bloomfield, S.F. 1996. Chlorine and iodine formulations, p. 133–158. In J. M. Ascenzi (ed.), Handbook of disinfectants and antiseptics. Marcel Dekker, Inc., New York, N.Y.
• Chevalier, J., Pages J.M., Eyraud, A., ve Mallea, M., 2000. Membrane permeability modifications are involved in antibiotic resistance in Klebsiella pneumoniae. Biochemical and Biophysical Research Communications, 274, 496-499.
• Chong, Z., Woo, W., Chng, S., 2015. Osmoporin OmpC forms a complex with MlaA to maintain outer membrane lipid asymmetry in Escherichia coli. Molecular Microbiology 98, 1133
• Darcan, C., 1999. Deniz suyunda dezenfektan özellikli kimyasal maddelerin Escherichia coli ML30 ve Salmonella typhimurium LT2 bakterilerinin protein sentezine olan etkileri. Yüksek Lisans Tezi, Ondokuz Mayıs Üniversitesi, Fen Bilimlerti Enstitüsü, Samsun, s.71 .
• Darcan, C., Özkanca, R., Şahin, N., Işık, K., ve Kariptaş, E., 2001. Dezenfektan özellikteki bazı kimyasal maddelerin deniz suyundaki Escherichia coli ML30 ve Salmonella typhimurium LT2’nin dış membran protein sentezine etkisi. Biyoteknoloji (KÜKEM) Dergisi, 25 (2), 57-66.
• Darcan, C. 2005. Karadeniz Suyunda pH, Osmolarite ve Açlık Stresinin E. coli’nin Dış Membran Porin Protein Sentez Düzeyine Etkisinin Araştırılması. Ondokuz Mayıs Üniversitesi, Fen Bilimleri enstitüsü, Doktora Tezi. Samsun, 185 s.
• Darcan, C., 2012 Expression of OmpC and OmpF porin proteins and survival of Escherichia coli under photooxidative stress in Black Sea water. Aquatic Biology 17 (2), 97-105.
• De Mot, R. ve Van Der Leyden, J., 1991. Purification of a root adhesive outer membrane protein of root-colonizing Pseudomonas fluerescens. FEMS Microbiology Letters, 81, 323-328.
• Delihas, N., 1995. Egulation of gene expression by trans-encoded antisense RNA. Molecular Microbiology, 15, 411-414.
• Dorman, C.J., Jay, C.D. ve Free, A., 1999. Domain organization and oligomerization among H-NS-like nucleoid-associated proteins in bacteria. Trend in Microbiology, 7(3), 124- 128.
• Du, Y., Lv, X.T., Wu, Q.Y., Zhang, D.Y., Zhou, Y.T., Peng, L., Hu, H.Y. 2017. Formation and control of disinfection byproducts and toxicity during reclaimed water chlorination: A review. Journal of Environmental Science (China), 58, 51-63.
• El-Falaha, B.M.A., Russell, A.D., ve Furr, J.R., 1985. Effect of chlorhexidine diacetate and benzalkonium chloride on the viability of wild-type and envelope mutants of Escherichia coli and Pseudomonas aeruginosa. Letters in Appllied Microbiology, 1: 21–24.
• Erickson, M.C., Liao, J.Y., Webb, C.C., Habteselassie, M.Y., Cannon, J.L., 2018. Efficacy of chlorine as a disinfecting agent on produce-harvesting gloves contaminated with E. coli O157:H7 or Salmonella. Food Control, 86, 257-265.
• Faber, F., Egli, T. ve Harder, W., 1993. Transient repression of the synthesis of OmpF and aspartate transcarbamoylase in Escherichia coli K12 as a response to pollutant stress. FEMS Microbiology Letters, 111, 189-196.
• Ferenci, T., 1999. Regulation by nutrient limitation. Current Opinion in Microbiology 2, 208-213.
• Ghai, I. ve Ghai, S., 2017. Exploring bacterial outer membrane barrier to combat bad bugs. Infect Drug Resistance, 10, 261–273.
• Heyde, M. ve Portalier, R., 1987. Regulation of major outer membrane porin proteins of Escherichia coli K12 by pH. Molecular and General Genetics, 208, 511-517.
• Heyde, M., Laloil, P. ve Portalier, R., 2000. Involvement of carbon source and acetyl phosphate in the external-pH-dependent expression of porin genes in E. coli. Journal of Bacteriology, 182 (1), 198-202.
• Imlay, J., 2008. Cellular defenses against superoxide and hydrogen peroxide. Annual Review Biochemistry, 77, 755-776.
• Koebnik, R., Locher, K.P. ve Van Gelder, P., 2000. Structure and function of bacterial outer membrane proteins: barrels in a nutshell. Molecular Microbiology, 37 (2), 239-253.
• Kumar, A., Bhandari, A. ve Krishnaswamy, S., 2015. Sequence and structural perspectives of bacterial β-stranded porins. Protein Peptid Letters, 22, 8-22.
• Liu, X. ve Ferenci, T., 1998. Regulation of porin mediated outer membrane permeability by nutrient limitation in Escherichia coli. Journal of Bacteriology, 180 (15), 3917-3922.
• Liu, X. ve Ferenci, T., 2001. An analysis of multifactorial influences on the transcriptional control of ompF and ompC porin expression under nutrient limitation. Microbiology, 147, 2981-2989.
• Low, A. S., MacKenzle F. M., Gould, I. M. ve Booth, I. R., 2001. Protected environments allow parallel evolution of a bacterial pathogen in a patient subjected to long term antibiotic therapy. Molecular Microbiology, 42 (3), 619-630.
• Martinez Flores, I., Cano, R., Bustamante, V.H., Calva, E., ve Puente, J.L., 1999. The ompB opreon partially determines differential expression of OmpC in Salmonella typhi and Escherichia coli. Journal of Bacteriology, 181 (2), 556-562.
• Massicotte, R., Mafu, A.A., Ahmad, D., Deshaies, F., Pichette, G. ve Belhumeur. P., 2017. Comparison between flow cytometry and traditional culture methods for efficacy assessment of six disinfectant agents against nosocomial bacterial Species. Frontier Microbiology, 8, 112.
• Mcdonnell, G. ve Russell, A.D., 1999. Antiseptics and Disinfectants: Activity, Action, and Resistance. Clinical Microbiology Reviews, 12(1), 147–179.
• McKenna, S.M. ve Davies, K.J.A. 1988. The inhibition of bacterial growth by hypochlorous acid. Biochemistry Journal, 254, 685–692.
• Miller, H.J., 1992. A Short course in bacterial genetics, Cold Spring Harbor Laboratory Press, USA, p 456.
• Morales, E.H., Calderón, I.L., Collao, B., Gil, F., Porwollik, S., McClelland, M., ve Saavedra, C.P., 2012. Hypochlorous acid and hydrogen peroxide-induced negative regulation of S..enterica serovar typhimurium ompW by the response regulator ArcA. BMC Microbiology, 12, 63-74.
• Negrete, A., Shiloach, J., 2017. Improving E. coli growth performance by manipulating small RNA expression. Microbial Cell Factories, 16(1), 198
• Nikaido, H. ve Vaara, M., 1985. Molecular basis of bacterial outer membrane permeability. Microbiological Reviews, 49 (1), 1-32.
• Nitzan, Y., Deutsch, B. ve Peshatnikov, I., 2002. Diffusion of β-Lactam antibiotics through oligomeric or monomeric porin channels of some gram-negative bacteria. Current Microbiology, 45, 446-455.
• Ofori, I., Maddila, S., Lin, J. ve Jonnalagadda, S.B., 2017. Chlorine dioxide oxidation of Escherichia coli in water – A study of the disinfection kinetics and mechanism. Journal of Environmental Science and Health, Part A Toxic/Hazardous Substances and Environmental Engineering, 52 (7), 598-606.
• Orruno, M., Parada, C., Kaberdin, V.R., ve Arana, I., 2017. Survival of Escherichia coli under Nutrient-Deprived Conditions: Effect on Cell Envelope Subproteome. Escherichia coli - Recent Advances on Physiology, Pathogenesis and Biotechnological Applications" Chapter 20. Book edited by Amidou Samie, ISBN 978-953-51-3330-8.
• Özkanca, R., 1993. Survival and physiological status of E. coli in lake water under different nutrient conditions, Ph D., Deparment of Biological Sciences, University of Warwick, England, p. 297.
• Rosner, J.L., Chai, T.J., ve Foulds J., 1991. Regulation of OmpF porin expression by salicylate in Escherichia coli. Journal of Bacteriology, 173 (18), 5631-5638.
• Russell, A.D., ve Gould, G.W., 1988. Resistance of Enterobacteriaceae to preservatives and disinfectants. Journal of Applied Bacteriology, 65, 167S–195S.
• Russell, A. D., and J. R. Furr. 1986. Susceptibility of some porin- and lipopolysaccharide-deficient strains of Escherichia coli to some antiseptics and disinfectants. Journal of Hospital Infection, 8, 47–56.
• Sato, M., Machida, K., Arıkado, E., Saito, H., Kakegawa T., et al., 2000. Expression of outer membrane proteins in Escherichia coli growing at acid pH. Applied Environmental Microbiology, 66 (3), 943-947.
• Schulz, G.E., 2002. The structure of bacterial outer membrane proteins. Biochimica et Biophysica Acta, 1565, 308-317.
• Sheinfeld, R., Smith, M.S., ve Valdes, S., 2017. Sodium Dodecyl Sulfate-Eethylenediaminetetraacetic Acid Sensitive Phenotype Associated with ompC Deficient Escherichia coli Strains is Observed Primarily in Cells Growing in Stationary Phase and Less So in Cells Growing in Log Phase. Journal of Experimental Microbiology and Immunology (JEMI) 21; 123-126.
• Shih, K.L. ve Lederberg, J. 1976. Effects of chloramine on Bacillus subtilis deoxyribonucleic acid. Journal of Bacteriology, 125, 934–945.
• Sidorova, O.V., Khomenko, V.A., Portnyagina, O.Y., Likhatskaya, G.N., Vakorina, T.I., Kim, N.Y., Chistyulin, D.K., Solov'eva, T.F., Novikova, O.D., 2014. Mutant OmpF porins of Yersinia pseudotuberculosis with deletions of external loops: structure-functional and immunochemical properties. Biochemical and Biophysical Research Communications, 445(2), 428-432.
• Sleator, R.D. ve Hill, C., 2001. Bacterial osmoadaptation: the role of osmolytes in bacterial stress and virulence. FEMS Microbiology Reviews, 26, 49-71.
• Song, W., Bajaj, H., Nasrallah, C., Jiang, H., Winterhalter, M., Colletier, J.P., et al., 2015. Understanding voltage gating of Providencia stuartii porins at atomic level. PLoS Computational Biology, 11(5), e1004255.