کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1986636 | 1540256 | 2014 | 7 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Kinetics and conformational stability studies of recombinant leucine aminopeptidase
ترجمه فارسی عنوان
سینتیک و بررسی پایداری سازگاری آمینوپپتیداز لوسین نوترکیب
دانلود مقاله + سفارش ترجمه
دانلود مقاله ISI انگلیسی
رایگان برای ایرانیان
کلمات کلیدی
آمینوپپتیداز لوسین بازآفرینی، پارامترهای جنبشی ثبات سازگاری،
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
چکیده انگلیسی
Leucine aminopeptidase from Vibrio proteolyticus is a broad specificity N-terminal aminopeptidase that is widely used in pharmaceutical processes where the removal of N-terminal residues in recombinant proteins is required. We previously reported the expression of a heterologous construction of the mature protein fused to a 6-histidine tag that presents a reasonable refolding rate for its use at industrial level. Here, we investigate this recombinant leucine aminopeptidase (rLAP) to explain the gain of activity observed when incubated at 37 °C after its production. Unfolding transitions of rLAP as a function of urea concentration were monitored by circular dichroism (CD) and fluorescence (FL) spectroscopy exhibiting single transitions by both techniques. Free energy change for unfolding measured by CD and FL spectroscopy are 2.8 ± 0.4 and 3.7 ± 0.4 kcal molâ1, respectively. Thermal stability conformation of rLAP is 2.6 ± 0.1 and 6.1 kcal molâ1 for CD and Nano-Differential Scanning Calorimetry (Nano-DSC), respectively. Enzyme activity was assessed with l-leucine-p-nitroanilide (L-pNA) as substrate. The catalytic efficiency was 3.87 ± 0.10 minâ1 μMâ1 at 37 °C and pH 8.0. Kinetic and conformation studies show differences between the enzyme native and rLAP; however rLAP is selective and specific to remove N-terminal groups from amino acids.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: International Journal of Biological Macromolecules - Volume 64, March 2014, Pages 306-312
Journal: International Journal of Biological Macromolecules - Volume 64, March 2014, Pages 306-312
نویسندگان
Ana V. Hernández-Moreno, Francisco Villaseñor, Emilio Medina-Rivero, Néstor O. Pérez, Luis F. Flores-Ortiz, Gloria Saab-Rincón, Gabriel Luna-Bárcenas,