Centaurin beta 2 Overexpression Lysate

Western Blot: Centaurin beta 2 Overexpression Lysate (Adult Normal) [NBL1-09095] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Centaurin beta 2.

• Reactivity: Hu
• Applications: WB

Centaurin beta 2 Overexpression Lysate Summary

• Description:

  Centaurin beta 2 Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC209171
  
  Accession#: NM_012287
  
  Protein Tag: C-MYC/DDK
  
  You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
• Gene: ACAP2

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.
  
  Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.
• Theoretical MW: 87.8 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for Centaurin beta 2 Overexpression Lysate

• Arf GAP with coiled coil, ANK repeat and PH domains 2
• ArfGAP with coiled-coil, ankyrin repeat and PH domains 2
• centaurin, beta 2
• Centaurin-beta-2
• CENTB2centaurin-beta-2
• CNT-B2
• KIAA0041arf-GAP with coiled-coil, ANK repeat and PH domain-containing protein 2

Background

Centaurin beta 1 and Centaurin beta 2 are recruited to platelet-derived growth factor-induced dorsal membrane ruffles in NIH 3T3 mouse fibroblasts, and overexpression inhibited ruffle formation. In vitro, Centaurin beta 1 and Centaurin beta 2 preferred ARF6 as substrate rather than ARF1 or ARF5, and the GAP activity of both Centaurins was dependent upon phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). The isolated PH domain of mouse Centaurin beta 2 exhibits moderate affinity for PtdIns(3,5)P2, but it does not bind any other phosphoinositides tested, including PtdIns(4,5)P2. Mutation of a highly conserved arginine in both Centaurins result in lack of ARF-GAP activity. Overexpression in HeLa cells of either Centaurin blocks the formation of ARF6-dependent protrusions. Both Centaurins are also recruited to peripheral, tubular membranes, where activation of ARF6 occurs and allows membrane recycling back to the plasma membrane.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Bioinformatics

• Gene Symbol: ACAP2