skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: A Link between Integral Membrane Protein Expression and Simulated Integration Efficiency

Authors:
; ; ; ; ; ; ; ;
Publication Date:
Sponsoring Org.:
USDOE
OSTI Identifier:
1378281
Resource Type:
Journal Article: Published Article
Journal Name:
Cell Reports
Additional Journal Information:
Journal Volume: 16; Journal Issue: 8; Related Information: CHORUS Timestamp: 2017-09-02 09:31:12; Journal ID: ISSN 2211-1247
Publisher:
Elsevier
Country of Publication:
Netherlands
Language:
English

Citation Formats

Marshall, Stephen S., Niesen, Michiel J. M., Müller, Axel, Tiemann, Katrin, Saladi, Shyam M., Galimidi, Rachel P., Zhang, Bin, Clemons, Jr., William M., and Miller, III, Thomas F. A Link between Integral Membrane Protein Expression and Simulated Integration Efficiency. Netherlands: N. p., 2016. Web. doi:10.1016/j.celrep.2016.07.042.
Marshall, Stephen S., Niesen, Michiel J. M., Müller, Axel, Tiemann, Katrin, Saladi, Shyam M., Galimidi, Rachel P., Zhang, Bin, Clemons, Jr., William M., & Miller, III, Thomas F. A Link between Integral Membrane Protein Expression and Simulated Integration Efficiency. Netherlands. doi:10.1016/j.celrep.2016.07.042.
Marshall, Stephen S., Niesen, Michiel J. M., Müller, Axel, Tiemann, Katrin, Saladi, Shyam M., Galimidi, Rachel P., Zhang, Bin, Clemons, Jr., William M., and Miller, III, Thomas F. Mon . "A Link between Integral Membrane Protein Expression and Simulated Integration Efficiency". Netherlands. doi:10.1016/j.celrep.2016.07.042.
@article{osti_1378281,
title = {A Link between Integral Membrane Protein Expression and Simulated Integration Efficiency},
author = {Marshall, Stephen S. and Niesen, Michiel J. M. and Müller, Axel and Tiemann, Katrin and Saladi, Shyam M. and Galimidi, Rachel P. and Zhang, Bin and Clemons, Jr., William M. and Miller, III, Thomas F.},
abstractNote = {},
doi = {10.1016/j.celrep.2016.07.042},
journal = {Cell Reports},
number = 8,
volume = 16,
place = {Netherlands},
year = {Mon Aug 01 00:00:00 EDT 2016},
month = {Mon Aug 01 00:00:00 EDT 2016}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record at 10.1016/j.celrep.2016.07.042

Citation Metrics:
Cited by: 3works
Citation information provided by
Web of Science

Save / Share:
  • Specific contacts between the lac repressor and operator have been explored using 5-bromodeoxyuridine-substituted DNA. Substitution of BrdU for single thymidine positions in a synthetic 40-base pair operator provides substrate for ultraviolet irradiation; upon irradiation, strand scission occurs at the BrdU residues. When bound, lac repressor protein provides protection against UV-induced breakage depending on the nature of the sites and type of interaction. We have confirmed 13 unique sites of inducer-sensitive protection along the operator sequence using this method compared to complete substitution with BrdU; differences were observed at two positions for singly substituted versus completely substituted DNAs. The ability ofmore » these photosensitive DNAs to form short range cross-links to bound protein has been used to determine the efficiency with which cross-linked protein-DNA complexes are generated at each individual site of BrdU substitution. Five sites of high efficiency cross-linking to the repressor protein have been identified. At one site, cross-linking without protection from strand scission was observed; this result suggests an unusual mechanism of strand scission and/or cross-linking at this site. Comparison of the UV protection results and the cross-linking data show that these processes provide complementary tools for identifying and analyzing individual protein-DNA contacts.« less
  • The human gene EPB72 coding for the band 7 integral membrane protein, a major protein of the erythrocyte membrane membrane, was isolated from a genomic DNA library and characterized. Spanning {approximately}30 kb, the human EPB72 gene comprises seven exons; intron sizes range from 970 to {approximately}11,200 bp. The first exon contains the 5{prime}-untranslated region (61 nucleotides) and the coding sequence for the N-terminal domain; the second exon encodes the hydrophobic domain, including flanking cysteine and lysine residues. Exon 7 contains the C-terminal portion and a 2-kb 3{prime}-untranslated region. The potential promoter region contains several consensus sequences for ubiquitous transcription factorsmore » (Sp1, AP1, AP2, CP1/2, NF{kappa}B, CREB, Ets-1, and CACC/GT-BF) and two imperfect sequences for erythroid factors (EKLF and GATA-1), in accordance with the ubiquitous distribution of the EPB72 mRNA in different cell types. No TATA box was apparent. An inverted Alu repeat element, flanked by nonamer direct repeats, was identified within the region -913/-620, relative to the cap site. Six additional Alu repeat elements, including one monomer and one trimer, were identified within the introns and the 3{prime}-untranslated region. Two polyadenylation signals in the 3{prime}-noncoding region of exon 7 enable the production of two mRNA species. 45 refs., 3 figs., 2 tabs.« less
  • Bacterial histidine kinases play an important role in the response to external stimuli. Structural studies of the histidine kinase transmembrane domain are challenging due to difficulties in protein expression and sample preparation. After carrying out expression screening of a series of histidine kinases, we investigated sample preparation methods for obtaining high quality samples of the periplasmic and transmembrane domain (PTD) of the bacterial histidine kinase SCO3062. Various sample conditions were tested for their ability to give homogeneous NMR spectra of the SCO3062 PTD with well-resolved resonances. Circular dichroism and 3D {sup 15}N-edited NOESY spectrum results demonstrate that the SCO3062 PTDmore » is predominantly {alpha}-helical. This method should be applicable to the NMR analysis of other transmembrane proteins.« less
  • Myristolyation of seven different {alpha} subunits of guanine nucleotide-binding regulatory proteins (G proteins) was examined by expressing these proteins in monkey kidney COS cells. Metabolic labeling studies of cells transfected with cytomegalovirus-based expression vectors indicated that ({sup 3}H)myristate was incorporated into {alpha}{sub i1}, {alpha}{sub i2}, {alpha}{sub i3}, {alpha}{sub 0}, and {alpha}{sub 1}, and {alpha}{sub z} but not {alpha}{sub s} subunits. The role of myristoylation in the association of {alpha} subunits with membranes was analyzed by site-directed mutagenesis and by substitution of myristate with a less hydrophobic analog, 10-(propoxy)decanoate (11-oxamyristate). Myristoylation of {alpha}{sub 0} was blocked when an alanine residue wasmore » substituted for its amino-terminal glycine, as was association of the protein with membranes. Substitution of the myristoyl group with 11-oxamyristate affected the cellular distribution of a subset of acylated {alpha} subunits. The results are consistent with a model wherein the hydrophobic interaction of myristate with the bilayer permits continued association of the protein with the plasma membrane when G-protein {alpha} subunits dissociated from {beta}{gamma}.« less
  • The Neisseria meningitidis outer membrane protein PorB was expressed in Escherichia coli and purified from inclusion bodies by denaturation in urea followed by refolding in buffered LDAO on a size-exclusion column. PorB has been crystallized in three different crystal forms: C222, R32 and P6{sub 3}. The C222 crystal form may contain either one or two PorB monomers in the asymmetric unit, while both the R32 and P6{sub 3} crystal forms contained one PorB monomer in the asymmetric unit. Of the three, the P6{sub 3} crystal form had the best diffraction quality, yielding data extending to 2.3 {angstrom} resolution.