Leptin-B Tilapia
Greater than 95.0% as determined by:
(a) Gel filtration analysis.
(b) Analysis by SDS-PAGE.
Description
Leptin-B Tilapia Recombinant produced in E.Coli is a single, non-glycosylated polypeptide chain containing 152 amino acids and having a molecular mass of 15,243 Dalton.
The Leptin-B Tilapia is purified by proprietary chromatographic techniques.
Product Specs
Leptin, a protein hormone primarily produced by adipose cells, plays a crucial role in energy homeostasis by regulating hunger. It exerts its effects by binding to nuclear receptors located in the hypothalamus, specifically the arcuate nucleus. Similar to insulin resistance observed in type II diabetes, obesity is characterized by reduced sensitivity to leptin, leading to an inability to sense satiety despite elevated energy stores and leptin levels. Tilapia (Oreochromis niloticus) possess full-length cDNA sequences encoding two leptin isoforms, tLepA and tLepB, as well as a leptin receptor, tLepR. The tLepR cDNA, spanning 3423 bp, encodes a protein of 1140 amino acids containing all essential functional domains conserved among vertebrate leptin receptors. The tLepA and tLepB cDNAs, measuring 486 bp and 459 bp, respectively, encode proteins of 161 and 152 amino acids. Notably, the three-dimensional structures of tLepA and tLepB exhibit significant conservation with human leptin, comprising four alpha-helices.
Recombinant Tilapia Leptin-B, expressed in E. coli, is a single, non-glycosylated polypeptide chain consisting of 152 amino acids, with a molecular weight of 15,243 Daltons. The purification process involves proprietary chromatographic techniques.
Lyophilized from a concentrated solution (1 mg/mL) containing NaHCO3 at a 1:2 salt-to-protein ratio.
For reconstitution, it is advised to dissolve the lyophilized Tilapia Leptin-B in sterile water or 0.4% NaHCO3 adjusted to a pH of 8-9, at a concentration not less than 100 µg/mL. This solution can then be further diluted in other aqueous solutions as needed.
Lyophilized Tilapia Leptin-B, while stable at room temperature for up to 3 weeks, should be stored desiccated at or below -18°C. Upon reconstitution, it is recommended to store Tilapia Leptin-B at 4°C for 2-7 days. For extended storage, freezing below -18°C is advisable. The addition of a carrier protein such as 0.1% HSA or BSA is recommended for long-term storage. It is crucial to avoid repeated freeze-thaw cycles.
The purity is determined to be greater than 95.0% using the following methods:
(a) Gel filtration analysis.
(b) SDS-PAGE analysis.
Tilapia leptins have demonstrated biological activity by promoting the proliferation of BAF/3 cells stably transfected with the long form of the human leptin receptor. However, their potency is lower compared to mammalian leptin. Additionally, Tilapia leptins exhibited biological activity by inducing STAT-LUC activation in COS7 cells transfected with the Tilapia leptin receptor, but not in cells expressing the human leptin receptor. Notably, Tilapia Leptin A displayed higher activity than Tilapia Leptin B.
Protein quantification was performed using UV spectroscopy at 280 nm. An absorbance value of 0.19 for a 1 mg/mL solution of Leptin-B Tilapia at pH 8.0 was used as the extinction coefficient. This value was determined using the DNAman computer analysis program for protein sequences.
The first six N-terminal amino acids of recombinant Tilapia leptin B are Ala-Leu-Leu-Thr-Lys-Gly.
Product Science Overview
Leptin is a protein hormone primarily produced in adipose cells. It plays a crucial role in regulating energy homeostasis by inhibiting hunger, thus helping to maintain body weight. Leptin binds to nuclear receptors in the hypothalamus, particularly in the arcuate nucleus. In cases of obesity, there is often a decrease in sensitivity to leptin, leading to a failure to recognize satiety despite high levels of energy stores or leptin itself.
In tilapia (Oreochromis niloticus), two leptin sequences (tLepA and tLepB) and one leptin receptor sequence (tLepR) have been identified. The full-length cDNA of tLepR is 3423 base pairs long, encoding a protein of 1140 amino acids. This protein contains all the functionally important domains conserved among vertebrate leptin receptors. The cDNAs of tLepA and tLepB are 486 and 459 base pairs long, encoding proteins of 161 and 152 amino acids, respectively. The three-dimensional structures of tLepA and tLepB show strong conservation of tertiary structure with that of human leptin, comprising four helices .
Leptin-B Tilapia Recombinant is produced in Escherichia coli (E. coli) as a single, non-glycosylated polypeptide chain containing 152 amino acids and having a molecular mass of 15,243 Daltons. The recombinant protein is purified using proprietary chromatographic techniques .
Physical Appearance and Formulation
Leptin-B Tilapia Recombinant is available as a sterile, filtered, white lyophilized (freeze-dried) powder. It is lyophilized from a concentrated solution containing sodium bicarbonate (NaHCO3) at a 1:2 salt-to-protein ratio. For reconstitution, it is recommended to use sterile water or 0.4% NaHCO3 adjusted to pH 8-9, at a concentration of no less than 100 µg/ml, which can then be further diluted into other aqueous solutions .
Stability and Storage
The lyophilized form of Leptin-B Tilapia is stable at room temperature for up to three weeks. However, for long-term storage, it should be kept desiccated below -18°C. Upon reconstitution, it should be stored at 4°C for 2-7 days and for future use below -18°C. To ensure long-term stability, it is recommended to add a carrier protein such as 0.1% human serum albumin (HSA) or bovine serum albumin (BSA). Freeze-thaw cycles should be avoided .
Purity and Biological Activity
Leptin-B Tilapia Recombinant has a purity greater than 95%, as determined by gel filtration analysis and SDS-PAGE. The first six N-terminal amino acids of the recombinant protein are Ala-Leu-Leu-Thr-Lys-Gly. Biologically, tilapia leptins have been found to promote the proliferation of BAF/3 cells stably transfected with the long form of the human leptin receptor, although their activity is lower than that of mammalian leptin. Additionally, tilapia leptins are active in promoting STAT-LUC activation in COS7 cells transfected with the tilapia leptin receptor but not in cells transfected with the human leptin receptor. Among the tilapia leptins, Leptin A is more active than Leptin B .
