شناسایی یک پپتید جدید مشتق از لاکتوفرین جدا شده از شیر شتر با فعالیت ضد میکروبی بالقوه
محورهای موضوعی : میکروب شناسی
الناز خواجه
1
,
مجید جمشیدیان مجاور
2
,
محسن نعیمی پور
3
,
حمیدرضا فرزین
4
1 - کارشناسی ارشد میکروبیولوژی، دانشکده علوم پایه، گروه زیست‌شناسی، واحد سبزوار، دانشگاه آزاد اسلامی، سبزوار، ایران.
2 - استادیار، موسسه تحقیقات واکسن و سرم‌سازی رازی، سازمان تحقیقات، آموزش و ترویج کشاورزی، مشهد، ایران
3 - استادیار، مرکز تحقیقات سلولی و مولکولی، دانشگاه علوم پزشکی سبزوار، سبزوار، ایران
4 - استادیار، موسسه تحقیقات واکسن و سرم‌سازی رازی، سازمان تحقیقات، آموزش و ترویج کشاورزی، شعبه مشهد، مشهد ایران
کلید واژه: شیر شتر, لاکتوفرین, تریپسین, پپتید ضد میکروب,
چکیده مقاله :
هدف: افزایش مقاومت میکروارگانیسم‌ها به آنتی بیوتیک‌های رایج، منجر به یافتن ترکیبات ضد میکروبی جدید شده است. پپتیدهای ضد میکروبی یکی از گزینه‌های مناسب برای جایگزینی آنتی بیوتیک‌های موجود می‌باشند. شیر حاوی تعداد زیادی پروتئین می‌باشد که برخی از آنها مانند لاکتوفرین، دارای فعالیت ضد باکتری است. هدف پژوهش حاضر بررسی فعالیت ضد باکتریایی یک پپتید مشتق شده از لاکتوفرین جدا شده از شیر شتر در برابر استافیلوکوکوس اورئوس، استرپتوکوکوس پیوژنز، سودوموناس آئروژینوزا و آسینتوباکتربومانی می‌باشد. مواد و روش‌ها: در پژوهش حاضر با استفاده از ابزارهای بیوانفورماتیکی به شناسایی پپتیدهای ضد باکتری در لاکتوفرین شیر پرداخته شد. پپتید تریپسین سنتز گردید. سپس با روش MTT خاصیت سمی پپتید بر رده سلولی بررسی شد. همچنین خاصیت ضد باکتری تریپسین بر چهار باکتری بیماری‌‌زای استافیلوکوکوس اورئوس، سودوموناس آئروژینوزا، آسینتوباکتربومانی و استرپتوکوکوس پیوژنز بررسی گردید. نتایج: نتایج نشان داد که پپتیدها بر رده سلولی مورد آزمایش اثر کشندگی نداشتند. نتایج MIC پپتید تریپسین برای باکتری‌های استافیلوکوکوس اورئوس، سودوموناس آئروژینوزا، آسینتوباکتربومانی و استرپتوکوکوس پیوژنز به ترتیب 81/7، 62/15، 125 و 250 بود. نتیجه‌گیری: پپتیدهای ضد میکروبی به علت برخورداری از خصوصیات و ویژگی‌‌های مناسب مانند کشندگی سریع، طیف وسیع فعالیت و همچنین پیشرفت نادر موارد ابتلاء به مقاومت دارویی، در دهه‌های اخیر بسیار مورد توجه قرار گرفته‌‌اند. با توجه نتایج مشاهده شده در این پژوهش، خاصیت آنتی‌‌باکتریال ترکیبات جدا شده از این تحقیق می‌‌توانند گزینه مناسبی برای جایگزینی با آنتی‌‌بیوتیک‌های متداول باشند.
Introduction: The increasing microbial resistance to existing antibiotics has increased the interest in novel antimicrobial compounds. Antimicrobial peptides (AMPs) represent an attractive alternative to classical antibiotics.Milk contains a lot of proteins, some of which have received a lot of attention, such as lactoferrin, which has antibacterial activity. The aim of this study was to investigate the antibacterial activity of a lactoferrin-derived peptide isolated from camel milk against Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa and, Acinetobacter baumannii. Materials and methods: In the present study, antibacterial peptides in milk lactoferrin were identified using bioinformatics tools. Trypsin I peptide was synthesized. Then, the toxicity of the peptide on the cell line was investigated by the MTT method. The antibacterial properties of trypsin I was evaluated on four pathogenic bacteria, Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii and, Streptococcus pyogenes. Results: The results showed that the peptides had no lethal effect on the cell line tested. The MIC results of trypsin peptide for Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii and, Streptococcus pyogenes were 7.81, 15.62, 125 and 250, respectively. Conclusion: Antimicrobial peptides have received much attention in recent decades due to their appropriate properties and characteristics such as rapid lethality, a wide range of activity and, also the rare development of cases of drug resistance. According to the observed results of this study, the antibacterial properties of the compounds isolated from this study can be a good alternative to replacement with common antibiotics.
Playne MJ, Bennett LE, Smithers GW. Functional dairy foods and ingredients. Australian Journal of Dairy Technology. 2003; 58(3): 242-64.
Miller GD, Jarvis JK & McBean LD. Handbook of dairy foods and nutrition. CRC press; 2006: 15.
Legrand D & Mazurier J. A critical review of the roles of host lactoferrin in immunity. Biometals. 2010; 23(3): 365-76.
Feng X, Liu C, Guo J, Bi C, Cheng B, Li Z, Shan A & Li Z. Expression and purification of an antimicrobial peptide, bovine lactoferricin derivative LfcinB-W10 in Escherichia coli. Current microbiology. 2010; 60(3): 179-84.
Metz‐Boutigue MH, Jollés J, Mazurier J, Schoentgen F, Legrand D, Spik G, Montreuil J & Jollès P. Human lactotransferrin: Amino acid sequence and structural comparisons with other transferrins. European Journal of Biochemistry. 1984; 145(3): 659-76.
Baker EN. Structure and reactivity of transferrins. Advances in inorganic chemistry. 1994; 41: 389-464.
Yen CC, Shen CJ, Hsu WH, Chang YH, Lin HT, Chen HL & Chen CM. Lactoferrin: An iron-binding antimicrobial protein against Escherichia coli infection. Biometals. 2011; 24(4): 585-94.
Expósito IL & Recio I. Antibacterial activity of peptides and folding variants from milk proteins. International Dairy Journal. 2006; 16(11): 1294-305.
Arnold RR, Brewer M & Gauthier JJ. Bactericidal activity of human lactoferrin: sensitivity of a variety of microorganisms. Infection and immunity. 1980; 28(3): 893-8.
Drago-Serrano ME, De La Garza-Amaya M, Luna JS & Campos-Rodríguez R. Lactoferrin-lipopolysaccharide (LPS) binding as key to antibacterial and antiendotoxic effects. International immunopharmacology. 2012; 12(1): 1-9.
Baker EN, Baker HM & Kidd RD. Lactoferrin and transferrin: functional variations on a common structural framework. Biochemistry and Cell Biology. 2002; 80(1): 27-34.
Kim WS, Shimazaki KI & Tamura T. Expression of bovine lactoferrin C-lobe in Rhodococcus erythropolis and its purification and characterization. Bioscience, biotechnology, and biochemistry. 2006; 70(11): 2641-5.
Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. Journal of immunological methods. 1983; 65 (1-2): 55-63.
Frija LM, Ntungwe E, Sitarek P, Andrade JM, Toma M, Śliwiński T, Cabral LS, Cristiano ML, Rijo P & Pombeiro AJ. In Vitro Assessment of Antimicrobial, Antioxidant, and Cytotoxic Properties of Saccharin-Tetrazolyl and-Thiadiazolyl Derivatives: The Simple Dependence of the pH Value on Antimicrobial Activity. Pharmaceuticals. 2019; 12(4): 167.
Kadoughani Sani S, Jamshidian-Mojaver M, Farzin H & Amiri M. Investigation of the effect of fungal and copper nanocomplexes on bacte-ria causing nosocomial infections. New Findings in Veterinary Microbiology. 2020; 3(1): 39-46.
Wallmann J, Böttner A, Goossens L, Hafez HM, Hartmann K, Kaspar H, Kehrenberg C, Kietzmann M, Klarmann D, Klein G & Krabisch P. Results of an interlaboratory test on antimicrobial susceptibility testing of bacteria from animals by broth microdilution. International journal of antimicrobial agents. 2006; 27(6): 482-90.
Balouiri M, Sadiki M & Ibnsouda SK. Methods for in vitro evaluating antimicrobial activity: A review. Journal of pharmaceutical analysis. 2016; 6(2): 71-9.
Ciccaglione AF, Di Giulio M, Di Lodovico S, Di Campli E, Cellini L & Marzio L. Bovine lactoferrin enhances the efficacy of levofloxacin-based triple therapy as first-line treatment of Helicobacter pylori infection: an in vitro and in vivo study. Journal of Antimicrobial Chemotherapy. 2019; 74(4): 1069-77.
Keil B. Proteolysis Data Bank: Specificity of alpha-chymotrypsin from computation of protein cleavages. Protein sequences & data analysis. 1987; 1(1): 13.
Gasteiger E, Hoogland C, Gattiker A, Wilkins MR, Appel RD & Bairoch A. Protein identification and analysis tools on the ExPASy server. In The proteomics protocols handbook. 2005; 74(4): 65-77.
De Gobba C, Espejo-Carpio FJ, Skibsted LH & Otte J. Antioxidant peptides from goat milk protein fractions hydrolysed by two commercial proteases. International Dairy Journal. 2014; 39(1): 28-40.
Sharbafi R, Moradian F, Rafiei AR. Evaluation of Antimicrobial Activity of Lactoferrin against P. Aeruginosa and E. Coli Growth. Journal of Babol University of Medical Sciences. 2016; 18(7): 19-23.
Taheri A, Farvin KS, Jacobsen C & Baron CP. Antioxidant activities and functional properties of protein and peptide fractions isolated from salted herring brine. Food chemistry. 2014; 142: 318-26.
Habibi Najafi MB, Tanhaeiyan A, Rahnama P & Azghandi M. Study on antibacterial activity and cytotoxicity of recombinant peptide, Lasioglossin ɪɪɪ, against foodborne pathogens. Agricultural Biotechnology Journal. 2018; 10(3): 31-44.
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Playne MJ, Bennett LE, Smithers GW. Functional dairy foods and ingredients. Australian Journal of Dairy Technology. 2003; 58(3): 242-64.
Miller GD, Jarvis JK & McBean LD. Handbook of dairy foods and nutrition. CRC press; 2006: 15.
Legrand D & Mazurier J. A critical review of the roles of host lactoferrin in immunity. Biometals. 2010; 23(3): 365-76.
Feng X, Liu C, Guo J, Bi C, Cheng B, Li Z, Shan A & Li Z. Expression and purification of an antimicrobial peptide, bovine lactoferricin derivative LfcinB-W10 in Escherichia coli. Current microbiology. 2010; 60(3): 179-84.
Metz‐Boutigue MH, Jollés J, Mazurier J, Schoentgen F, Legrand D, Spik G, Montreuil J & Jollès P. Human lactotransferrin: Amino acid sequence and structural comparisons with other transferrins. European Journal of Biochemistry. 1984; 145(3): 659-76.
Baker EN. Structure and reactivity of transferrins. Advances in inorganic chemistry. 1994; 41: 389-464.
Yen CC, Shen CJ, Hsu WH, Chang YH, Lin HT, Chen HL & Chen CM. Lactoferrin: An iron-binding antimicrobial protein against Escherichia coli infection. Biometals. 2011; 24(4): 585-94.
Expósito IL & Recio I. Antibacterial activity of peptides and folding variants from milk proteins. International Dairy Journal. 2006; 16(11): 1294-305.
Arnold RR, Brewer M & Gauthier JJ. Bactericidal activity of human lactoferrin: sensitivity of a variety of microorganisms. Infection and immunity. 1980; 28(3): 893-8.
Drago-Serrano ME, De La Garza-Amaya M, Luna JS & Campos-Rodríguez R. Lactoferrin-lipopolysaccharide (LPS) binding as key to antibacterial and antiendotoxic effects. International immunopharmacology. 2012; 12(1): 1-9.
Baker EN, Baker HM & Kidd RD. Lactoferrin and transferrin: functional variations on a common structural framework. Biochemistry and Cell Biology. 2002; 80(1): 27-34.
Kim WS, Shimazaki KI & Tamura T. Expression of bovine lactoferrin C-lobe in Rhodococcus erythropolis and its purification and characterization. Bioscience, biotechnology, and biochemistry. 2006; 70(11): 2641-5.
Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. Journal of immunological methods. 1983; 65 (1-2): 55-63.
Frija LM, Ntungwe E, Sitarek P, Andrade JM, Toma M, Śliwiński T, Cabral LS, Cristiano ML, Rijo P & Pombeiro AJ. In Vitro Assessment of Antimicrobial, Antioxidant, and Cytotoxic Properties of Saccharin-Tetrazolyl and-Thiadiazolyl Derivatives: The Simple Dependence of the pH Value on Antimicrobial Activity. Pharmaceuticals. 2019; 12(4): 167.
Kadoughani Sani S, Jamshidian-Mojaver M, Farzin H & Amiri M. Investigation of the effect of fungal and copper nanocomplexes on bacte-ria causing nosocomial infections. New Findings in Veterinary Microbiology. 2020; 3(1): 39-46.
Wallmann J, Böttner A, Goossens L, Hafez HM, Hartmann K, Kaspar H, Kehrenberg C, Kietzmann M, Klarmann D, Klein G & Krabisch P. Results of an interlaboratory test on antimicrobial susceptibility testing of bacteria from animals by broth microdilution. International journal of antimicrobial agents. 2006; 27(6): 482-90.
Balouiri M, Sadiki M & Ibnsouda SK. Methods for in vitro evaluating antimicrobial activity: A review. Journal of pharmaceutical analysis. 2016; 6(2): 71-9.
Ciccaglione AF, Di Giulio M, Di Lodovico S, Di Campli E, Cellini L & Marzio L. Bovine lactoferrin enhances the efficacy of levofloxacin-based triple therapy as first-line treatment of Helicobacter pylori infection: an in vitro and in vivo study. Journal of Antimicrobial Chemotherapy. 2019; 74(4): 1069-77.
Keil B. Proteolysis Data Bank: Specificity of alpha-chymotrypsin from computation of protein cleavages. Protein sequences & data analysis. 1987; 1(1): 13.
Gasteiger E, Hoogland C, Gattiker A, Wilkins MR, Appel RD & Bairoch A. Protein identification and analysis tools on the ExPASy server. In The proteomics protocols handbook. 2005; 74(4): 65-77.
De Gobba C, Espejo-Carpio FJ, Skibsted LH & Otte J. Antioxidant peptides from goat milk protein fractions hydrolysed by two commercial proteases. International Dairy Journal. 2014; 39(1): 28-40.
Sharbafi R, Moradian F, Rafiei AR. Evaluation of Antimicrobial Activity of Lactoferrin against P. Aeruginosa and E. Coli Growth. Journal of Babol University of Medical Sciences. 2016; 18(7): 19-23.
Taheri A, Farvin KS, Jacobsen C & Baron CP. Antioxidant activities and functional properties of protein and peptide fractions isolated from salted herring brine. Food chemistry. 2014; 142: 318-26.
Habibi Najafi MB, Tanhaeiyan A, Rahnama P & Azghandi M. Study on antibacterial activity and cytotoxicity of recombinant peptide, Lasioglossin ɪɪɪ, against foodborne pathogens. Agricultural Biotechnology Journal. 2018; 10(3): 31-44.