Type
Recombinant protein
Description
Total 235 AA. MW: 26.6 kDa (calculated). UniProtKB acc.no. Q9Y2Q3 (Gly2-Leu226). N-terminal his-tag (10 extra AA). Protein identity confirmed by LC-MS/MS.
Amino Acid Sequence
MKHHHHHHASGPLPRTVELFYDVLSPYSWLGFEILCRYQNIWNINLQLRPSLITGIMKDSGNKPPGLLPRKGLYMANDLKLLRHHLQIPIHFPKDFLSVMLEKGSLSAMRFLTAVNLEHPEMLEKASRELWMRVWSRNEDITEPQSILAAAEKAGMSAEQAQGLLEKIATPKVKNQLKETTEAACRYGAFGLPITVAHVDGQTHMLFGSDRMELLAHLLGEKWMGPIPPAVNARL
Source
E. coli
Purity
Purity as determined by densitometric image analysis: >95 %
SDS-PAGE Gel
14 % SDS-PAGE separation of Human DsbA-L:
1. M.W. marker – 14, 21, 31, 45, 66, 97 kDa
2. Reduced and boiled sample, 2.5 μg/lane
3. Non-reduced and non-boiled sample, 2.5 μg/lane
Endotoxin
< 1.0 EU/μg
Formulation
Filtered (0.4 μm) and lyophilized in phosphate buffered saline pH 7.0 + 5%(w/v) trehalose.
Reconstitution
Add deionized water to prepare a working stock solution of approximately 0.5 mg/ml and let the lyophilized pellet dissolve completely. Filter sterilize your culture media/working solutions containing this non-sterile product before using in cell culture.
Applications
Western blotting, ELISA
Shipping
At ambient temperature. Upon receipt, store the product at the temperature recommended below.
Storage/Expiration
Store the lyophilized protein at –80 °C. Lyophilized protein remains stable until the expiry date when stored at –80 °C. Aliquot reconstituted protein to avoid repeated freezing/thawing cycles and store at –80 °C for long term storage. Reconstituted protein can be stored at 4 °C for a week.
Quality Control Test
BCA to determine quantity of the protein.
SDS PAGE to determine purity of the protein.
LAL to determine quantity of endotoxin.
Note
This product is intended for research use only.
Research topic
Diabetology - Other Relevant Products, Energy metabolism and body weight regulation
Summary
Disulfide-bond A oxidoreductase-like protein (DsbA-L) is the enzyme, which used to be called glutathione S-transferase kappa 1 (GSTK1) in humans and belongs to superfamily of enzymes glutathione S-transferases (GST), which are mainly known for cellular detoxification. GSTK1 enzyme is a homodimer, molecular weight of each subunit being 27 kDa, and shares high sequence and secondary structure homology with bacterial DsbA. Based on this finding, the enzyme GSTK1 was renamed to DsbA-L.
Bacterial DsbA catalyzes disulfide bond formation during folding of secreted proteins and therefore DsbA-L might also have the ability to catalyze disulfide bond formation during protein assembling. DsbA-L is expressed in a number of mouse tissues such as liver, kidney, pancreas, heart and lung; the highest DsbA-L expression was observed in the adipose tissue. It has been shown that DsbA-L expression in adipocytes is negatively correlated with obesity in mice and humans. DsbA-L in adipocytes plays a critical role in regulation of adiponectin secretion and multimerization to form high molecular weight (HMW) complex. This HMW form of adiponectin is the active form of the hormone and has a relevant role in enhancing insulin sensitivity and in protecting against diabetes. Impairment in adiponectin multimerization leads to defects in adiponectin secretion and function and is associated with diabetes. These findings suggest that increasing the ratio of the HMW form rather than the total levels of adiponectin might provide an effective alternative therapeutic strategy.
Transgenic mice overexpressing DsbA-L in fat exhibited increased levels of total and HMW adiponectin compared with wild type mice. These mice also displayed suppression of diet-induced obesity, insulin resistance and hepatic steatosis compared with wild type mice; thus, upregulation of DsbA-L may be a new therapeutic approach to treatment of obesity and associated metabolic disorders.
Originally, DsbA-L was identified as a mitochondrial enzyme but recent investigation provides evidence that DsbA-L is also localized in endoplasmic reticulum (ER). In obesity, increased demands on ER function (protein folding) lead to ER stress which in turn downregulates cellular levels and secretion of adiponectin in 3T3-L1 adipocytes. Experiments with 3T3-L1 adipocytes have shown that DsbA-L localization in ER is critical for its promoting effect on adiponectin biosynthesis in adipocytes and for suppressing the negative effects of ER stress.
