This Mouse TIM-1 / KIM-1 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of TIM-1 / KIM-1 protein (Cat: 50321-M16H) from the overexpression lysate was verified.
A DNA sequence encoding the extracellular domain of mouse HAVCR1 isoform b (NP_001160104.1) (Tyr 22-Thr 212) was fused with a polyhistidine tag at the C-terminus and the Fc region of human IgG1 at the N-terminus.
The recombinant mouse HAVCR1/Fc is a disulfide-linked homodimer. The reduced monomer comprises 438 amino acids and predicts a molecular mass of 49 kDa. As a result of glycosylation, the apparent molecular mass of HAVCR1/Fc monomer is approximately 80-85 kDa in SDS-PAGE under reducing conditions.
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.
HAV cellular receptor 1 (HAVCR1), also known as Kidney injury molecule 1 (KIM-1) and T cell immunoglobulinmucin 1 (TIM-1), is a type â… integral membrane glycoprotein. KIM-1 protein is widely expressed with highest levels in kidney and testis. It has been shown to play a major role as a human susceptibility gene for asthma, allergy and autoimmunity. IgA1lambda is a specific ligand of KIM-1 protein and that their association has a synergistic effect in virus-receptor interactions. KIM-1 involves in the pathogenesis of acute kidney injury. It had been confirmed that KIM-1 is a human urinary renal dysfunction biomarker. Moreover, KIM-1 protein is a novel regulatory molecule of flow-induced calcium signaling.
Tami C, et al. (2007) Immunoglobulin A (IgA) is a natural ligand of hepatitis A virus cellular receptor 1 (HAVCR1), and the association of IgA with HAVCR1 enhances virus-receptor interactions. J Virol. 81(7): 3437-46.
Rees AJ, et al. (2008) Kim-1/Tim-1: from biomarker to therapeutic target? Nephrol Dial Transplant. 23(11): 3394-6.
Chaturvedi S, et al. (2009) Assay validation for KIM-1: human urinary renal dysfunction biomarker. Int J Biol Sci. 5(2): 128-34.