This Human PCSK1 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of PCSK1 protein (Cat: 10129-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the human PCSK1 (NP_000430.3) (Met 1-Arg 617) was fused with a polyhistidine tag at the C-terminus.
The purified recombinant human PCSK1 consists of 518 amino acids after removal of the pro peptide due to autocatalytic processing, corresponding to the active form (aa 111-617), and has a predicted molecular mass of 57.4 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of the active rh PCSK1 is approximately 66 kDa due to glycosylation.
Human PCSK1 HEK293 Overexpression Lysate: 使用指南
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.
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.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
稳定性 & 储存条件
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.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human PCSK1 HEK293 Overexpression Lysate: 别称
Human BMIQ12 Overexpression Lysate; Human NEC1 Overexpression Lysate; Human PC1 Overexpression Lysate; Human PC3 Overexpression Lysate; Human SPC3 Overexpression Lysate
Neuroendocrine convertase 1, also known as Prohormone convertase 1, Proprotein convertase 1, PCSK1 and NEC1, is anenzyme which belongs to thepeptidase S8 family and Furin subfamily. PCSK1 is an enzyme that performs the proteolytic cleavage of prohormones to their intermediate (or sometimes completely cleaved) forms. It is present only in neuroendocrine cells such as brain, pituitary and adrenal, and most often cleaves after a pair of basic residues within prohormones but can occasionally cleave after a single arginine. It binds to a protein known as proSAAS, which also represents its endogenous inhibitor. PCSK1 is involved in the processing of hormone and other protein precursors at sites comprised of pairs of basic amino acid residues. PCSK1 substrates include POMC, renin, enkephalin, dynorphin, somatostatin and insulin. Defects in PCSK1 are the cause of proprotein convertase 1 deficiency (PC1 deficiency). PC1 deficiency is characterized by obesity, hypogonadism, hypoadrenalism, reactive hypoglycemia as well as marked small-intestinal absorptive dysfunction. It is due to impaired processing of prohormones.