کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8359469 | 1542294 | 2018 | 11 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Full-length myocilin protein is purified from mammalian cells as a dimer
ترجمه فارسی عنوان
پروتئین میوسیلین در تمام طول از سلول های پستاندار به عنوان یک دیمر پاک می شود
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کلمات کلیدی
oLFiOpMYOCOlfactomedin - اولفکتومدینBeta-mercaptoethanol - بتا موراپتواتانولfull-length - تمام طولProtein isolation - جداسازی پروتئینProtein purification - خالصسازی یا تخلیص پروتئینDimer - دیمرaqueous humor - طنز آبیIntraocular pressure - فشار داخل چشمmyocilin - میوسیلینwild-type - نوع وحشیMolecular weight - وزن مولکولی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
چکیده انگلیسی
Myocilin (MYOC) is a secreted protein found in human aqueous humor (AH) and mutations in the MYOC gene are the most common mutation observed in glaucoma patients. Human AH analyzed under non-reducing conditions suggests that MYOC is not normally found in a monomeric form, but rather is predominantly dimeric. Although MYOC was first reported almost 20 years ago, a technical challenge still faced by researchers is an inability to isolate full-length MYOC protein for experimental purposes. Herein we describe two methods by which to isolate sufficient quantities of human full-length MYOC protein from mammalian cells. One method involved identification of a cell line (HeLa S3) that would secrete full-length protein (15â¯mg/L) while the second method involved a purification approach from 293â¯cells requiring identification and modification of an internal MYOC cleavage site (Glu214/Leu215). MYOC protein yield from 293â¯cells was improved by mutation of two MYOC N-terminal cysteines (C47 and C61) to serines. Analytical size exclusion chromatography of our full-length MYOC protein purified from 293â¯cells indicated that it is predominantly dimeric and we propose a structure for the MYOC dimer. We hope that by providing methods to obtain MYOC protein, researchers will be able to utilize the protein to obtain new insights into MYOC biology. The ultimate goal of MYOC research is to better understand this target so we can help the patient that carries a MYOC mutation retain vision and maintain quality of life.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 147, July 2018, Pages 38-48
Journal: Protein Expression and Purification - Volume 147, July 2018, Pages 38-48
نویسندگان
Parvaneh Katoli, Adarsh Godbole, Michael J. Romanowski, Kirk Clark, Erik Meredith, Veronica Saenz-Vash, Y. Karen Wang, Nancy Lewicki, Andrew A. Nguyen, Jeffrey M. Lynch,