Purification and characterization of organic solvent tolerant extracellular lipase from bacterial isolate K5B4

العناوين الأخرى

تنقية و توصيف أنزيم الليبيز خارج الخلوي المحتمل للمذيبات العضوية و المفرز من العزلة البكتيرية k5b4

مقدم أطروحة جامعية

al-Rawashidah, Saddam Ata Allah Awdah

مشرف أطروحة جامعية

al-Khulayfat, Khalid Muhammad Awf

أعضاء اللجنة

al-Shakhanbah, Jumah Muti Muhammad
al-Shurafa, Khalid Yusuf Ahmad
al-Laymun, Muhammad Awdah

الجامعة

جامعة مؤتة

الكلية

كلية العلوم

القسم الأكاديمي

قسم الأحياء

دولة الجامعة

الأردن

الدرجة العلمية

ماجستير

تاريخ الدرجة العلمية

2016

الملخص الإنجليزي

In this project the extracellular lipase enzyme produced by Acinetobacter sp.

K5b4 was purified to homogeneity using ultrafiltration (Amicon tube, cutoff 30 KDa) followed by gel filtration chromatography on Sephadex G-50.

Enzyme activities were detected early in fractions number 8, 9, 10 and 11.

The SDS-PAGE analysis of the collected fractions (8, 9, 10 and 11) indicated that there was one single protein band with relative molecular mass estimated to be 133 KDa.

The lipase enzyme was purified 8.99 fold with recovery of 30% (fraction 10).

The Km and Vmax value of the purified enzyme for pNPL hydrolysis were calculated to be 4.0 mM and 73.53 nmol/ml/min, respectively.

The pure enzyme was highly activated in the presence of 20, 40 and 60% (v/v) methanol, DMSO and acetone meanwhile, ethanol, acetonitrile and propanol caused reduction in the enzyme activity.

Maximum enzyme activity was recorded at optimum pH of 7.0 and 27 oC with pH stability ranging from slight acidic to neutral pH (6.5-7.0) and temperature lower than 30oC.

No effects to slight enhancement in the enzyme activity were observed with the mineral salts KCl, BaCl2 and MgCl2 while CoCl2, ZnCl2, MnCl2and CuCl2 caused slight to medium reduction of 10-20% in the enzyme activity.

When CaCl2 was added to the reaction mixture a remarkable increase in the enzyme activity (36%) was observed.

No inhibitory effects were observed with 1.0 and 5.0 mM of 2-Mercaptoethanol and EDTA.

Similarly, SDS at 1.0 mM does not affect the enzyme activity while high reduction (80%) was observed at 5.0 mM SDS concentration.

The enzyme was active against p-nitrophenyl esters of C8, C12 and C16 with highest preference to the medium carbon chain p-nitrophenyl caprylate (C8).

The fact that the enzyme displays distinct stability in the presence of methanol, DMSO and acetone suggests that this lipase is suitable as biocatalyst in organic synthesis where such hydrophilic organic solvents are used as a reaction media

التخصصات الرئيسية

الأحياء

عدد الصفحات

44

قائمة المحتويات

• APA
• MLA
• AMA

لغة النص

الإنجليزية

نوع البيانات

رسائل جامعية

رقم السجل

BIM-731541