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@ARTICLE{Wittlich:60123,
      author       = {Wittlich, M. and Wiesehan, K. and Koenig, B. W. and
                      Willbold, D.},
      title        = {{E}xpression, purification and membrane reconstitution of a
                      {CD}4 fragment comprising the transmembrane and cyctoplasmic
                      domains of the receptor},
      journal      = {Protein expression and purification},
      volume       = {55},
      issn         = {1046-5928},
      address      = {Orlando, Fla.},
      publisher    = {Academic Press},
      reportid     = {PreJuSER-60123},
      pages        = {198 - 207},
      year         = {2007},
      note         = {Record converted from VDB: 12.11.2012},
      abstract     = {The transmembrane glycoprotein CD4 plays a prominent role
                      in the adaptive immune response. CD4 is displayed primarily
                      on the surface of T helper cells, but also on subsets of
                      memory and regulatory T lymphocytes, macrophages, and
                      dendritic cells. Binding of the lymphocyte specific tyrosine
                      kinase p56(lck) to the cytoplasmic domain of CD4 is crucial
                      for antigen receptor-mediated signal transduction. The human
                      immunodeficiency virus (HIV) utilizes CD4 as the main
                      receptor for T cell invasion. The virus has developed
                      multiple strategies for down-regulation of CD4 in infected
                      cells. Physical interactions of viral proteins VpU and Nef
                      with the cytoplasmic tail of CD4 initiate a cascade of
                      events leading to degradation of CD4. Here we report
                      heterologous expression and purification of a CD4 fragment
                      comprising the transmembrane and cytoplasmic domains of
                      human CD4. A synthetic gene encoding CD4 amino acid residues
                      372-433 and a protease cleavage site was cloned into the
                      pTKK19xb/ub plasmid. The CD4 fragment was expressed in
                      Escherichia coli C43(DE3) cells as a ubiquitin fusion with
                      an N-terminal His-tag, isolated, released by PreScission
                      proteolytic cleavage, and purified to homogeneity.
                      Incorporation of the recombinant CD4 fragment in lipid
                      membranes and physical interaction with the cytoplasmic
                      domain of VpU was demonstrated by centrifugation assays
                      followed by reversed phase chromatographic analysis of the
                      composition of the proteoliposomes. A high resolution NMR
                      spectrum of uniformly (15)N-labeled CD4 peptide in membrane
                      simulating micelles proves the possibility of solution NMR
                      studies of this CD4 fragment and of its molecular
                      complexes.},
      keywords     = {Amino Acid Sequence / Antigens, CD4: biosynthesis /
                      Antigens, CD4: genetics / Antigens, CD4: isolation $\&$
                      purification / Cell Membrane: chemistry / Escherichia coli:
                      genetics / Genes, Synthetic / Humans / Liposomes: chemistry
                      / Molecular Sequence Data / Nuclear Magnetic Resonance,
                      Biomolecular / Protein Structure, Tertiary / Recombinant
                      Proteins: biosynthesis / Recombinant Proteins: chemistry /
                      Recombinant Proteins: isolation $\&$ purification / Viral
                      Regulatory and Accessory Proteins: chemistry / Antigens, CD4
                      (NLM Chemicals) / Liposomes (NLM Chemicals) / Recombinant
                      Proteins (NLM Chemicals) / Viral Regulatory and Accessory
                      Proteins (NLM Chemicals) / J (WoSType)},
      cin          = {INB-2 / JARA-SIM},
      ddc          = {570},
      cid          = {I:(DE-Juel1)VDB805 / I:(DE-Juel1)VDB1045},
      pnm          = {Funktion und Dysfunktion des Nervensystems},
      pid          = {G:(DE-Juel1)FUEK409},
      shelfmark    = {Biochemical Research Methods / Biochemistry $\&$ Molecular
                      Biology / Biotechnology $\&$ Applied Microbiology},
      typ          = {PUB:(DE-HGF)16},
      pubmed       = {pmid:17613246},
      UT           = {WOS:000249631800023},
      doi          = {10.1016/j.pep.2007.05.007},
      url          = {https://juser.fz-juelich.de/record/60123},
}