Recombinant Human Amyloid Beta (A4) Precursor Protein, His-tagged

• Cat.No.: APP-051H
• Product Overview: Recombinant humanAPP protein,fused to His-tag at N-terminus, was expressed in E. coli andpurified by using conventional chromatography.

Specification

• Species: Human
• Source: E.coli
• Tag: His
• Description: APP iscleaved by secretases to form a number of peptides. Some of these peptidesare secreted and can bind to the acetyltransferase complex APBB1/TIP60 topromote transcriptional activation, while others form the protein basis of theamyloid plaques found in the brains of patients with Alzheimer disease.Mutations in this gene have been implicated in autosomal dominant Alzheimerdisease and cerebroarterial amyloidosis (cerebral amyloidangiopathy).
• Form: Liquid. 20mMTris-HCl buffer (pH8.0) containing 20% glycerol, 0.1M NaCl, 1mM DTT.
• Molecular Weight: 34.7 kDa (308confirmed by MALDI-TOF
• Purity: > 85% by SDS-PAGE
• Concentration: 1 mg/ml(determined by Bradford assay)
• Sequences of aminoacids: MRGSHHHHHHGMASMTGGQQ MGRDLYDDDD KDRWGSLEVP TDGNAGLLAE PQIAMFCGRL NMHMNVQNGKWDSDPSGTKT CIDTKEGILQ YCQEVYPELQ ITNVVEANQP VTIQNWCKRG RKQCKTHPHF VIPYRCLVGEFVSDALLVPD KCKFLHQERM DVCETHLHWH TVAKETCSEK STNLHDYGML LPCGIDKFRG VEFVCCPLAEESDNVDSADA EEDDSDVWWG GADTDYADGS EDKVVEVAEE EEVAEVEEEE ADDDEDDEDG DEVEEEAEEPYEEATERTTS IATTTTTTTE SVEEVVRE
• Storage: Can bestored at +4°C short term (1-2 weeks). For long term storage, aliquot andstore at -20°C or -70°C. Avoid repeated freezing and thawing cycles.
• OfficialSymbol: APP
• Pathway: Activated TLR4 signalling;Alzheimer"s disease; Formation of Platelet plug; GPCR ligand binding;Glypican 1 network; Hemostasis; Innate Immunity Signaling; PlateletActivation; Platelet degranulation; Signaling by GPCR; Signaling in Immunesystem; TRAF6 mediated NF-kB activation; Toll Receptor Cascades; Toll LikeReceptor 2 Cascade; Toll Like Receptor 3 (TLR3) Cascade

Gene Information

• Gene Name: APP amyloid beta (A4) precursorprotein [ Homo sapiens ]
• Synonyms: APP; amyloid beta (A4) precursor protein;AAA; AD1; PN2; ABPP; APPI; CVAP; ABETA; CTFgamma; amyloid beta A4 protein;PN-II; preA4; protease nexin-II; OTTHUMP00000096095; OTTHUMP00000096096;OTTHUMP00000096097; OTTHUMP00000096098; OTTHUMP00000225525; peptidasenexin-II; beta-amyloid peptide; alzheimer disease amyloid protein; cerebralvascular amyloid peptide; Alzheimer disease; human mRNA for amyloid A4precursor of Alzheimer"s disease; A4
• Gene ID: 351
• mRNA Refseq: NM_000484
• Protein Refseq: NP_000475
• MIM: 104760
• UniProt ID: P05067
• Chromosome Location: 21q21.2

Citation

HIV-1 Tat Interacts with and Regulates the Localization and Processing of Amyloid Precursor Protein

Journal: PLoS ONE    PubMed ID: 24312169    Data: 2022/11/28

Authors: Jiyoung Kim, Jee-Hyun Yoon, Ashok Chauhan

Article Snippet:Beads were washed three times with PBS, bound proteins were eluted by boiling in SDS-PAGE buffer and analyzed by 8% SDS-PAGE followed by western blotting with anti-APP (22C11) antibody.Beads were washed three times with PBS, bound proteins were eluted by boiling in SDS-PAGE buffer and analyzed by 8% SDS-PAGE followed by western blotting with anti-APP (22C11) antibody.. Purified recombinant APP (cat. no APP-526H, Creative BioMart, NY, USA) was resuspended in PBS supplemented with 1% NP-40 to yield a final concentration of 1 ng/μl, and 500 μl was incubated with GST- or GST-Tat-coated beads for 3 hours at 4°C.. The beads were washed three times with PBS supplemented with 1% NP-40.The beads were washed three times with PBS supplemented with 1% NP-40.

( A ) GST and GST-Tat were purified on glutathione-Sepharose beads. The beads were boiled to elute the bound proteins, which were then run on a 12% SDS-PAGE gel and stained with Coomassie brilliant blue (left panel). GST pulldown assay with SK-N-MC neuroblastoma cell lysates shows a strong interaction between APP and GST-Tat (right panel). SK-N-MC neuroblastoma cell extracts incubated with GST- or GST-Tat-coated beads for 3 hours. The beads were washed three times with PBS and the eluted proteins were analyzed by western blotting with an anti-APP antibody (22C11). ( B ) Coimmunoprecipitation of Tat and APP in HEK 293FT cells transfected with Tat and/or Myc-tagged APP695 vectors. Proteins were precipitated with anti-Tat or anti-APP (6E10) antibodies and immunoblotted with anti-Tat or anti-Myc antibodies. ( C ) Coimmunoprecipitation of U-87 MG cell lysates transduced with mock, Lenti-Tat, or Lenti-mTat virus. APP was precipitated with an APP antibody (6E10), and the precipitate was analyzed by SDS-PAGE followed by Western blotting with anti-APP (22C11) or anti-Tat antibodies. Reciprocally, Tat was precipitated with anti-Tat antibody, and the precipitate was analyzed by SDS-PAGE followed by Western blotting with anti-APP (A8717) or anti-Tat antibody. ( D ) Purified recombinant APP interacts with GST-Tat. Purified recombinant APP (500 ng) was incubated with GST- or GST-Tat-coated beads and the eluted proteins were analyzed by western blotting with an APP antibody. A large amount of recombinant APP bound to the GST-Tat beads. ( E ) Tat interacts strongly with APP. SK-N-MC neurobalstoma cell lysates were incubated with GST, GST-Tat, or GST-Tat beads, and washed three times in buffer containing 137, 200, 300, 400 or 500 mM NaCl. APP remained associated with GST-Tat under high-salt conditions. ( F ) The cysteine-rich domain of Tat is important for association with APP. Deletion mutants were produced as GST-fusion proteins and subjected to GST-pulldown assays with SK-N-MC cell lysates. L, load; B; bound.