Human MAP1D Baculovirus-Insect cells Overexpression Lysate

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Human MAP1D Baculovirus-Insect cells Overexpression Lysate: Product Information

Product Description
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.
Expression Host
Baculovirus-Insect cells
Species
Human
Sequence Information
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.
Molecule Mass
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: Usage Guide

Preparation Method
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.
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.
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.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
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.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human MAP1D Baculovirus-Insect cells Overexpression Lysate: Alternative Names

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

MAP1D Background Information

Methionine aminopeptidase 1D, also known as MAP1D, is a member of thepeptidase 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-independant 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.
Full Name
methionyl aminopeptidase type 1D (mitochondrial)
References
  • 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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