Human Carboxypeptidase E/CPE HEK293 Overexpression Lysate

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Human Carboxypeptidase E/CPE HEK293 Overexpression Lysate: 製品の情報

製品の説明
This Human Carboxypeptidase E/CPE overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Carboxypeptidase E/CPE protein (Cat: 10069-H02H) from the overexpression lysate was verified.
発現ホスト
HEK293 Cells
Human
タンパク質構築情報
A DNA sequence encoding the human carboxypeptidase E (CPE) precursor (NP_001864.1) (Met 1-Ser 453) was expressed with the C-terminal fused Fc region of human IgG1.
分子量
The recombinant human CPE/Fc is a disulfide-linked homodimeric protein. The reduced monomer consists of 666 amino acids and predicts a molecular mass of 74.6 kDa. As a result of glycosylation, the apparent molecular mass of rhCPE/Fc monomer is approximately 85-90 kDa in SDS-PAGE under reducing conditions.

Human Carboxypeptidase E/CPE HEK293 Overexpression Lysate: 用法

調製方法
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
溶解バッファー
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
おすすめの用法
1.  Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2.  Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
バッファー
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
安定性 & 保存条件
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
アプリケーション
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Carboxypeptidase E/CPE 背景情報

Carboxypeptidase E (CPE), also known as Carboxypeptidase H, is a peripheral membrane protein and a zinc metallocarboxypeptidase, and the conversion of proCPE into CPE occurs primarily in secretory vesicles. The active form of CPE cleaves C-terminal amino acid residues of the peptide, and is thus involved in the biosynthesis of peptide hormones and neurotransmitters including insulin, enkephalin, etc. The enzymatic activity is enhanced by millimolar concentrations of Co2+. It has also been proposed that membrane-associated carboxypeptidase E acts as a sorting receptor for targeting regulated secretory proteins which are mostly prohormones and neuropeptides in the trans-Golgi network of the pituitary and in secretory granules into the secretory pathway.Its interaction with glycosphingolipid-cholesterol rafts at the TGN facilitates the targeting. Mutations in this gene are implicated in type II diabetes due to impaired glucose clearance and insulin resistance.
完全な名称
carboxypeptidase E
参考文献
  • Manser, E. et al., 1990, Biochem. J. 267: 517-525.
  • Cool, D.R. et al., 1997, Cell. 88: 73-83.
  • Song, L. and Fricker, L. 1995, J. Neurochem. 65: 444-453.
  • Dhanvantari,S. et al., 2000, J. Biol. Chem. 275: 29887-29893.
  • Jeffrey, K.D. et al., 2008, Proc. Natl. Acad. Sci. U.S.A. 105: 8452-8457
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