کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
7022528 | 1456055 | 2013 | 6 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
One-step-purification of penicillin G amidase from cell lysate using ion-exchange membrane adsorbers
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
مهندسی شیمی
تصفیه و جداسازی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
This study describes the purification of penicillin G amidase (PGA) by ion exchange membrane adsorbers in a one-step-process. Preliminary experiments with high-throughput screening devices in microliter scale (8-strip modules) were performed to find suitable purification strategy and appropriate ion exchange ligands as well as basic process conditions for binding and elution. Best purification results were achieved by strong cation-exchange (S-) membrane adsorbers loaded with 2Â ml/min enzyme solution at pH 6.0 and eluted at pH 6.0 with 0.05Â M NaCl, which led to a high yield of bound PGA (98%) without any visible remains of host cell proteins and with a residual enzyme activity of 80-85%. The binding of PGA to the S-membrane was further investigated in an upscaling to milliliter scale with LP15 modules and breakthrough curves were determined by varying the flow rates: the PGA-binding to S-membrane adsorbers is independent of the flow rate. Dynamic binding capacities were estimated to be 0.69Â mg PGA/cm2 (25.5Â mg/ml) for 10% breakthrough respectively 0.95Â mg/cm2 (35.2Â mg/ml) for 100% breakthrough. Finally, real cell lysate samples from Escherichia coli culture containing PGA were processed under the found optimal conditions. Despite exceeded loading PGA was isolated from this complex mixture successfully fourfold concentrated and with a purification factor of 101.3 and a resulting specific activity of 4.97Â U/mg.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Membrane Science - Volume 444, 1 October 2013, Pages 359-364
Journal: Journal of Membrane Science - Volume 444, 1 October 2013, Pages 359-364
نویسندگان
Andrea Mönster, Louis Villain, Thomas Scheper, Sascha Beutel,