This Human CHST15 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of CHST15 protein (Cat: 11033-H07H) from the overexpression lysate was verified.
A DNA sequence encoding the human BRAG (NP_056976.2) extracellular domain (Ser 99-Thr 561) was expressed, with a polyhistidine tag at the N-terminus.
The recombinant human BRAG consists of 479 amino acids and predictes a molecular mass of 56 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rh BRAG is approximately 70-80 kDa due to glycosylation.
Human CHST15 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 CHST15 HEK293 Overexpression Lysate: 别称
Human BRAG Overexpression Lysate; Human GALNAC4S-6ST Overexpression Lysate
Carbohydrate sulfotransferase 15, also known as N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase, GalNAc4S-6ST, B-cell RAG-associated gene protein, CHST15 and BRAG, is a single-pass type II membrane protein which belongs to thesulfotransferase 1 family. CHST15 / BRAG is expressed in B-cell-enriched tissues but not in fetal or adult thymus. It is expressed in fetal and adult spleen, lymph node, tonsil, bone marrow and peripheral leukocytes. It is not expressed in T-cells. In pro-B, pre-B, and mature B-cell lines, it colocalizes with RAG1. CHST15 / BRAG is a sulfotransferase that transfers sulfate from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to the C-6 hydroxyl group of the GalNAc 4-sulfate residue of chondroitin sulfate A and forms chondroitin sulfate E containing GlcA-GalNAc(4,6-SO4) repeating units. It also transfers sulfate to a unique non-reducing terminal sequence, GalNAc(4SO4)-GlcA(2SO4)-GalNAc(6SO4), to yield a highly sulfated structure similar to the structure found in thrombomodulin chondroitin sulfate. CHST15 / BRAG may also act as a B-cell receptor involved in BCR ligation-mediated early activation that mediate regulatory signals key to B-cell development and / or regulation of B-cell-specific RAG expression.
Verkoczy L.K., et al.,(1998), hBRAG, a novel B cell lineage cDNA encoding a type II transmembrane glycoprotein potentially involved in the regulation of recombination activating gene 1 (RAG1). Eur. J. Immunol. 28:2839-2853.
Yuki M., et al., (2000), Structure, expression and mutational analysis of the hBRAG gene on 10q in the frequently deleted region in human endometrial cancer.Oncol. Rep. 7:1339-1342.
Ohtake S., et al.,(2001), Human N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase cDNA is related to human B cell recombination activating gene-associated gene.J. Biol. Chem. 276:43894-43900.