Certificate of Analysis and Data Sheet

Source:
E.Coli

Catalog No.
CYT-356

Background:

Leptin inhibits food intake and stimulates energy expenditure. Leptin also has thermogenic actions and regulates enzymes of fatty acid oxidation. Severe hereditary obesity in rodents and humans is caused by defects in leptin production .In addition to its critical role in the physiologic regulation of body weight leptin has a variety of other physiologic and pathologic functions resembling those of cytokines . These functions include the regulation of hematopoiesis , angiogenesis , wound healing , inflammation , and immune responses.

Description :

Recombinant ovine leptin, one polypeptide chain containing 146 amino and additional Ala at N-terminus acids and having a molecular mass of ~ 16 kDa, oLEP was mutated, resulting in L39A/D40A/F41A mutant that was purified by proprietary chromatographic techniques.

Physical Appearance:

Sterile Filtered White lyophilized (freeze-dried) powder.

Formulation:

The protein was lyophilized from a concentrated (0.65mg/ml) solution with 0.003mM NaHCO3.

Solubility:

It is recommended to reconstitute the lyophilized ovine LEPTIN mutant in sterile water or sterile 0.4% NaHCO3 adjusted to pH 8, not less than 100ug/ml, which can then be further diluted to other aqueous solutions.

Stability:

Lyophilized ovine LEP mutant although stable at room temperature for several weeks, should be stored desiccated below -18⁰C. Upon reconstitution at > 0.1 oLEP mutant mg/ml and up to 2 mM and filter sterilization oLEP mutant can be stored at 4⁰C or even room temperature for several weeks making it suitable for long term infusion studies using osmotic pumps. At lower concentration addition of a carrier protein (0.1% HSA or BSA) is suggested.

Please prevent freeze-thaw cycles.

Purity:

Greater than 98.0% as determined by:
(a) Gel filtration analysis.
(b) Analysis by reducing and non-reducing SDS-PAGE gel.

Amino acid sequence:

The sequence of the first five N-terminal amino acids was determined and was found to be Ala-Val-Pro-Ile-Arg

Dimers and aggregates:

The purified ovine LEP triple antagonist (16K) consists of > 95% monomers as determined by gel-filtration chromatography.

Biological Activity:

ProSpec旧 oLEP triple antagonist is capable of inhibiting leptin-induced proliferation of BAF/3 cells stably transfected with the long form of ovine leptin receptor. It also inhibits various leptin effects in several in vitro bioassays.

Endotoxin:

Less than 0.1 ng/ug (IEU/ug) of oLEP triple antagonist

Protein content:

Protein quantitation was carried out by UV spectroscopy at 280 nm using the absorbency value of 0.21 as the extinction coefficient for a 0.1% (1mg/ml) solution at pH 8.0. This value is calculated by the PC GENE computer analysis program of protein sequences (IntelliGenetics).

Usage:

Prospec's products are furnished for LABORATORY RESEARCH USE ONLY. The product may not be used as drugs, agricultural or pesticidal products, food additives or household chemicals.

Latest Publications:

1. Identification of the hydrophobic strand in the A-B loop of leptin as major binding site III: implications for large-scale preparation of potent recombinant human and ovine leptin antagonists.
Biochem J 2005 Oct 15;391(Pt 2):221-30
2. Chronic administration of recombinant ovine leptin in growing beef heifers: effects on secretion of LH, metabolic hormones, and timing of puberty.
J Anim Sci 2004 Oct;82(10):2930-6
3. Overexpression of ovine leptin in Pichia pastoris: physiological yeast response to leptin production and characterization of the recombinant hormone.
Yeast 2004 Feb;21(3):249-63
4. The late gestation increase in circulating ACTH and cortisol in the fetal sheep is suppressed by intracerebroventricular infusion of recombinant ovine leptin.
J Endocrinol 2002 Aug;174(2):259-66
5. Central infusion of recombinant ovine leptin normalizes plasma insulin and stimulates a novel hypersec retion of luteinizing hormone after short-term fasting in mature beef cows.
Biol Reprod 2002 May;66(5):1555-61
6. Effect of intravenous infusion of recombinant ovine leptin on feed intake and serum concentrations of GH, LH, insulin, IGF-1, cortisol, and thyroxine in growing prepubertal ewe lambs.
Domest Anim Endocrinol 2002 Apr;22(2):103-12