Isolement, identification et criblage d’actinomycètes à activité chitinolytique à partir du sol de la région de Laghouat: Optimisation de la production de chitinase par Streptomyces griseorubens C9 par la méthode des surfaces de réponse

الموضوعات

علوم بحتة وطبيعية

روابط وملفات

التعريف والنوع

رقم الوثيقة

fd874386-b2e0-4b16-a939-dde73a7dffb8

رقم العقد

0

نوع الوسائط

Crawler

نوع المحتوى

الرسائل العلمية

صيغة المصدر

رسائل دكتوراة

نوع الملف

pdf text

أسماء الملفات

796084_1.pdf

بيانات النشر

ألقاب المؤلفين

[{"name_ar":"Mahmoud Kitouni","title_ar":"اشراف","title_en":"Supervision"},{"name_ar":"Meriem Gasmi","title_ar":"اعداد","title_en":"Preparation"}]

اللغة

French

المصدر والدورية

اسم المصدر

Isolement, identification et criblage d’actinomycètes à activité chitinolytique à partir du sol de la région de Laghouat: Optimisation de la production de chitinase par Streptomyces griseorubens C9 par la méthode des surfaces de réponse

المحتوى والصفحات

عدد الصفحات

0

ترجمة الملخص

With at least 10 gigatons synthesized and degraded each year in the biosphere, chitin is the secondmost represented polysaccharide in nature after cellulose. It is a crystalline polysaccharide consistingof long linear chains containing more than 1000 units of N-acetyl-β-D-glucosamine linked by β-1,4glycoside bonds. Chitin degradation is performed by chitinases (E.C. 3.2.1.14), that hydrolyze ß-1,4bonds releasing compounds easily metabolized by many microbes. Production of microbial chitinasesreceives worldwide attention in both industrial and scientific communities, because of its wideapplications spectrum. Chitinases are used in biochemistry, food processing, and various chemicalindustries due their antimicrobial, anticholesterol, and antitumor activities. They are also used inwaste-water treatment, dietary fiber and in agriculture to inhibit plant pathogens. The physiologicalfunction of chitinases depends on their origin. In bacteria, chitinases play a trophic role by degradingchitin, as source of both carbon and nitrogen. Microorganisms particularly Streptomyces are efficientchitin hydrolysers. Chitinases have been identified in several Streptomyces spp. It is induced by chitinand repressed by readily utilizable carbon sources such as N-acetyl-glucosamine and glucose. In thisthesis, we chose to study actinobacteria isolated from a semi-arid soil of the Algerian desert in order toidentify strains with a high chitinolytic and antifungal potential. We isolated 40 phenotypicallydifferent actinobacteria. First, we carried out a functional study by screening these isolates on aminimal medium supplemented with colloidal chitin (CCM) to characterize their chitinolytic activitiesunder extreme temperature conditions (30, 35, 40, 45 and 50 °C) and pH (5, 7 and 9). In addition, ataxonomic study was conducted by combining a BOX-PCR approach and sequencing 16S rRNA of 20strains. This identification showed that 10 of these isolates belonged to the genus of Streptomyces, twoto the genus of Micromonospora and 08 potential new bacterial species. Also, the search forantimicrobial activity against 06 test bacteria and two phytopathogenic fungi was carried out, using anovel glucose-inducing / repressing chitinase approach in the antifungal test, to prove that onlychitinases can inhibit these phytopathogenic fungi. One strain (C31) was able to reduce fungal growthonly after induction of chitinase by colloidal chitin. The second part of this work aimed to usestatistical Plackett–Burman and Box–Wilson response surface methodology to optimize the mediumcomponents and, thus, improve chitinase production by Streptomyces griseorubens C9. First, syrup ofdate, colloidal chitin, yeast extract and K2HPO4, KH2PO4 were proved to have significant effects onchitinase activity using the Plackett–Burman design. Then, an optimal medium was obtained by aBox–Wilson factorial design of response surface methodology in liquid culture. Maximum chitinaseproduction (0.902 U / ml) was predicted in medium containing 2% colloidal chitin, 0.47% syrup ofdate, 0.25 g/l yeast extract and 1.81 g/l K2HPO4, KH2PO4.This was confirmed by Erlenmeyer cultureswhere the Production of chitinase by S. griseorubens C9 reached 1.53 U/ml, the yield of enzymeproduction was increased 26.38% after optimization by the response surface methodology. Finally,purification and partial characterization of the enzyme produced in the optimized medium wasperformed. 3 major purification steps were done and the enzyme reached a purification fold of 23.24; ayield of 29.7% and a specific activity equal to 2.393 U/mg. The purified chitinase activity hadoptimum catalytic activity at pH 5.0 and 50 °C. It retained more than 60% stability at pH rangebetween 4.0 and 7.0 and remained stable at temperatures between 30 °C and 60 °C. Thus, S.griseorubens C9 could be a potent strain for industrial chitinase production, as well as inbiotechnology and bio-control fields.

إشراف وإعداد

الإشراف

Mahmoud Kitouni

الإعداد

Meriem Gasmi

الاقتباسات الببليوغرافية