X-ray diffraction analysis of cytochrome b5 reconstituted in egg phosphatidylcholine vesicles

Rzepecki, Leszek M.; Strittmatter, Philipp

doi:10.1016/s0006-3495(86)83712-2

Cited by 12 publications

(14 citation statements)

References 39 publications

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“…The anchoring C-terminal peptide of cytochrome bs, reconstituted in its "tightly bound" form into lipid vesicles, may form a bent helix inserted part of the way into the bilayer. The evidence, however, is somewhat controversial (see Gogol, Engelman & ZaccaL 1983;Takagaki et al, 1983;Gogol & Engelman, 1984;Rzepecki, Strittmatter & Herbette, 1986;Arin% Rzepecki & Strittmatter, 1987) and not certain to reflect the native topology. To our knowledge, the two models initially proposed for PLP are unique in suggesting that a polytopic (multispanning) membrane protein features polypeptide segments looping back in the middle of the bilayer (Stoffel et al, 1983(Stoffel et al, , 1984Laursen et al, 1984;Fig.…”

Section: Introductionmentioning

confidence: 99%

Major myelin proteolipid: The 4-α-helix topology

1991

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Several conflicting models have been proposed for the membrane arrangement of the major myelin proteolipid (PLP). We have compared features of the sequence of PLP with those of other eukaryotic integral membrane proteins, with the view of identifying the most likely transmembrane topology. A new, simple model is suggested, which features four hydrophobic alpha-helices spanning the whole thickness of the lipid bilayer. Its orientation may be such that both the N- and C-termini face the cytosol. None of the biochemical, biophysical or immunological experiments hitherto reported provides incontrovertible evidence against the model. The effect or absence thereof of various PLP mutations is discussed in the frame of the proposed 4-helix topology.

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Section: Introductionmentioning

confidence: 99%

Major myelin proteolipid: The 4-α-helix topology

1991

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“…Insight into the phase assignment for this particular reflection (quite important to the extent of bilayer asymmetry) was obtained by Patterson deconvolution ( Fig. 4; Skita et al, (Rzepecki et al, 1986). *Data from Rzepecki et al (1986) was d/2 shifted to obtain structure shown in manuscript.…”

Section: Fluorescence Anisotropymentioning

confidence: 99%

“…the phase set determined by Rzepecki et al (1986) and those derived by our evaluation of the structure factor plot (Fig. 3).…”

Section: CXmentioning

confidence: 99%

“…Brominated-lipid tryptophan fluorescence quenching experiments of Everett et al (1986) are also consistent with a cis orientation. To date, x-ray (Rzepecki et al, 1986) and neutron (Gogol et al, 1983, Gogol and Engelman, 1984) diffraction studies on membrane bound cytochrome b5 have been unatble to discriminate between these two NPP structural configurations. Low resolution (15 A) x-ray studies reveal an asymmetric bilayer structure which is consistent with NPP penetration to the bilayer center (Rzepecki et al, 1986).…”

Section: Introductionmentioning

confidence: 99%

“…To date, x-ray (Rzepecki et al, 1986) and neutron (Gogol et al, 1983, Gogol and Engelman, 1984) diffraction studies on membrane bound cytochrome b5 have been unatble to discriminate between these two NPP structural configurations. Low resolution (15 A) x-ray studies reveal an asymmetric bilayer structure which is consistent with NPP penetration to the bilayer center (Rzepecki et al, 1986). This model is qualitatively consistent with calorimetry and resonance energy transfer studies suggesting that only one monolayer is perturbed by asymmetric cytochrome b5 incorporation (Friere et al, 1983).…”

Section: Introductionmentioning

confidence: 99%

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Bilayer structure and physical dynamics of the cytochrome b5 dimyristoylphosphatidylcholine interaction

Chester

Skita

Young

et al. 1992

Biophysical Journal

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Cytochrome b5 is a microsomal membrane protein which provides reducing potential to delta 5-, delta 6-, and delta 9-fatty acid desaturases through its interaction with cytochrome b5 reductase. Low angle x-ray diffraction has been used to determine the structure of an asymmetrically reconstituted cytochrome b5:DMPC model membrane system. Differential scanning calorimetry and fluorescence anisotropy studies were performed to examine the bilayer physical dynamics of this reconstituted system. These latter studies allow us to constrain structural models to those which are consistent with physical dynamics data. Additionally, because the nonpolar peptide secondary structure remains unclear, we tested the sensitivity of our model to different nonpolar peptide domain configurations. In this modeling approach, the nonpolar peptide moiety was arranged in the membrane to meet such chemically determined criteria as protease susceptibility of carboxyl- and amino-termini, tyrosine availability for pH titration and tryptophan 109 location, et cetera. In these studies, we have obtained a reconstituted cytochrome b5:DMPC bilayer structure at approximately 6.3 A resolution and conclude that the nonpolar peptide does not penetrate beyond the bilayer midplane. Structural correlations with calorimetry, fluorescence anisotropy and acyl chain packing data suggest that asymmetric cytochrome b5 incorporation into the bilayer increases acyl chain order. Additionally, we suggest that the heme peptide:bilayer interaction facilitates a discreet heme peptide orientation which would be dependent upon phospholipid headgroup composition.

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Reconstitution and Physiological Protein Translocation Processes

Etemadi

1989

Subcellular Biochemistry

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FIGURE 1. Some models for membrane-protein interaction during translocation of secretory and! or transmembrane integral proteins are reproduced. In all schemes, unless otherwise stated, the lower side and the upper side represent the cytoplasmic (cis) and the extracytoplasmic (trans) compartment, respectively. In the loop model A (DiRienzo et al. 1978;Inouye and Halegoua 1980), positively charged amino acid(s) at the N terminus of the signal sequence react with negatively charged head groups of membrane phospholipids. The hydrophobic part of the signal sequence would interact with and loop back in the hydrophobic core of the membrane, bending being due to the presence of a glycine or proline residue in the hydrophobic segment of the signal. Chain elongation would continue until the cleavage site reaches the side remote from the cytoplasm where processing occurs. In model B (Garnier et at., 1980) the signal sequence as a double amphipathic structure inserts in the membrane. Its hydrophobic core has an a-helical or extended conformation and its N terminus is extracytoplasmic. Only after this insertion ribophorin (R) assembly around the signal peptide occurs and interaction of ribophorins, with the ribosomes, leads to the formation of a channel through which the protein is translocated. In the helical hairpin model, C (Engelman and Steitz, 1981), the signal sequence and the N-terminal segment of the mature part of a nascent secretory or transmembrane integral protein form a hydrophobic (H) and a polar (P) helix, respectively, which together exhibit a helical hairpin conformation. The "helical hairpin" is inserted into the membrane in such a way that the N terminus of the signal sequence is cytoplasmic, as in the loop model. Continuation of translation extrudes residues of the polar helix in the trans side, while

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X-ray diffraction analysis of cytochrome b5 reconstituted in egg phosphatidylcholine vesicles

Cited by 12 publications

References 39 publications

Major myelin proteolipid: The 4-α-helix topology

Major myelin proteolipid: The 4-α-helix topology

Bilayer structure and physical dynamics of the cytochrome b5 dimyristoylphosphatidylcholine interaction

Reconstitution and Physiological Protein Translocation Processes

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