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Human CROT (474 Leu/Val) Insect Cell Lysate (WB positive control)

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Human CROT Transfected / Overexpression Cell Lysate Product Information
Expressed Host:Baculovirus-Insect cells
Product Description:Baculovirus-Insect Cell lysate that Human CROT transfected / overexpressed for Western blot (WB) positive control. The whole cell lysate is provided in 1X Sample Buffer (1X modified RIPA buffer+1X SDS loading buffer).
Sequence information:A DNA sequence encoding the human CROT (Q9UKG9) (Met 1-Leu 612, 474 Leu/Val) was fused with a polyhistidine tag at the C-terminus.
Predicted N Terminal:Met 1
Molecule Mass:The recombinant human CROT consists of 623 amino acids and predicts a molecular mass of 71.5 kDa. The apparent molecular mass of rhCROT is approximately 65 kDa in SDS-PAGE under reducing conditions.
Human CROT Transfected / Overexpression Cell Lysate Usage Guide
Preparation Method:Cell lysate was prepared by homogenization 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.
Lysis Buffer: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.
Quality Control Testing:12.5% SDS-PAGE Stained with Coomassie Blue after protein purification.
Stability:Samples are stable for up to twelve months from date of receipt.
Recommend Usage: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. 3. Store the lyophilized cell lysate at 4℃. After re-dissolution, recommend to aliquot it into smaller quantities and store at -80℃.
Storage Buffer:1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Storage Instruction:Store at 4℃. After re-dissolution, aliquot and store at -80℃.
Application notes:Western blot (WB): Use at an assay dependent dilution.
Other Applications: Not tested.
Optimal dilutions/concentrations should be determined by the end user.
CROT Background

Carnitine octanoyltransferase (CROT or COT), also known as octanoyl-CoA: L-carnitine O-octanoyltransferase, medium-chain/long-chain carnitine acyltransferase, and carnitine medium-chain acyltransferase, is a carnitine acyltransferase belonging to the family of transferases, specifically those acyltransferases transferring groups other than aminoacyl groups that catalyzes the reversible transfer of fatty acyl groups between CoA and carnitine. Carnitine octanoyltransferase (CROT or COT) facilitate the transport of medium- and long-chain fatty acids through the peroxisomal and mitochondrial membranes. It is physiologically inhibited by malonyl-CoA. COT also has functions in efficiently converting one of the end products of the peroxisomal beta-oxidation of pristanic acid, 4, 8-dimethylnonanoyl-CoA, to its corresponding carnitine ester. 

Human CROT References
  • Ferdinandusse S, et al. (1999) Molecular cloning and expression of human carnitine octanoyltransferase: evidence for its role in the peroxisomal beta-oxidation of branched-chain fatty acids. Biochem Biophys Res Commun. 263 (1): 213-8.
  • Feike R, et al. (2000) Genomics of the Human Carnitine Acyltransferase Genes. Molecular Genetics and Metabolism. 71 (1-2): 139-53.
  • Montserrat Morillas, et al. (2002) Structural Model of a Malonyl-CoA-binding Site of Carnitine Octanoyltransferase and Carnitine Palmitoyltransferase I. The Journal of Biological Chemistry, 277: 11473-80.
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    Catalog: 11015-H08BL-300
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