Human MAP1D Baculovirus-Insect cells Overexpression Lysate

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Human MAP1D Baculovirus-Insect cells Overexpression Lysate: 产品信息

产品描述
This Human MAP1D overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of MAP1D protein (Cat: 10883-H08B) from the overexpression lysate was verified.
表达宿主
Baculovirus-Insect cells
种属
Human
蛋白构建信息
A DNA sequence encoding the human MAP1D (NP_954697.1) (Arg 44-Ala 335) was expressed, with an initial Met at the N-terminus and a polyhistidine tag at the C-terminus.
分子量
The secreted recombinant human MAP1D consists of 303 amino acids and predicts a molecular mass of 33.4 kDa. It migrates as an approximately 36 kDa band in SDS-PAGE under reducing conditions.

Human MAP1D Baculovirus-Insect cells Overexpression Lysate: 使用指南

制备方法
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.
裂解缓冲液
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.
使用建议
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.
缓冲液
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.
应用
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human MAP1D Baculovirus-Insect cells Overexpression Lysate: 别称

Human MAP 1D Overexpression Lysate; Human MAP1D Overexpression Lysate; Human MetAP 1D Overexpression Lysate; Human Metap1l Overexpression Lysate

MAP1D 背景信息

Methionine aminopeptidase 1D, also known as MAP1D, is a member of the peptidase M24A family. N-terminal methionine removal is an important cellular process required for proper biological activity, subcellular localization, and eventual degradation of many proteins. The enzymes that catalyze this reaction are called Methionine aminopeptidases (MAPs). MAP1D is overexpressed in colon cancer cell lines and colon tumors as compared to normal tissues (at protein level). Downregulation of MAP1D expression by shRNA in HCT-116 colon carcinoma cells reduces anchorage-independent growth in soft agar. MAP1D binds two cobalt ions per subunit. The true nature of the physiological cofactor is under debate. MAP1D is also active with zinc, manganese, or divalent ions. MAP1D removes the amino-terminal methionine from nascent proteins. It may also play an important role in colon tumorigenesis.
全称
methionyl aminopeptidase type 1D (mitochondrial)
参考文献
  • Serero A., et al.,(2003), An unusual peptide deformylase features in the human mitochondrial N-terminal methionine excision pathway. J. Biol. Chem. 278:52953-52963.
  • Leszczyniecka M., et al., (2006), MAP1D, a novel methionine aminopeptidase family member is overexpressed in colon cancer.Oncogene 25:3471-3478.
  • Vaca Jacome A.S., et al.,(2015), N-terminome analysis of the human mitochondrial proteome.Proteomics 15:2519-2524.
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