Transmembrane helices mediate the formation of a stable ternary complex of cyt b5 reductase, cyt b5, and SCD1

Mammalian cytochrome b5 (cyt b5) and cytochrome b5 reductase (b5R) are electron carrier proteins required for many membrane-embedded oxidoreductases. Both cyt b5 and b5R have a cytosolic domain anchored to the membrane by a single transmembrane helix (TM). It is not clear if b5R, cyt b5 and their partner oxidoreductases assemble as binary or ternary complexes. Here we show that b5R and cyt b5 form a stable binary complex, and that b5R, cyt b5 and a membrane-embedded oxidoreductase, stearoyl-CoA desaturase 1 (SCD1) form a stable ternary complex. The formation of the complexes significantly enhances electron transfer rates, and that the single TM of cyt b5 and b5R mediated assembly of the complexes. These results reveal a novel functional role of TMs in cyt b5 and b5R and suggest that an electron transport chain composed of a stable ternary complex may be a general feature in oxidoreductases that require the participation of cyt b5 and b5R.

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Refined structure of the FKBP12–rapamycin–FRB ternary complex at 2.2 Å resolution

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s0907444998014747 ◽

1999 ◽

Vol 55 (4) ◽

pp. 736-744 ◽

Cited By ~ 78

Author(s):

Jun Liang ◽

Jungwon Choi ◽

Jon Clardy

Keyword(s):

Cell Cycle ◽

Biological Activity ◽

High Resolution ◽

Protein Interactions ◽

Ternary Complex ◽

Binary Complex ◽

Fk506 Binding Protein ◽

Cycle Arrest ◽

Protein Partners

The structure of the FKBP12–rapamycin–FRB ternary complex has now been refined at 2.2 Å resolution. The cell-cycle arrest agent rapamycin binds FK506-binding protein (FKBP12) and the FKBP12–rapamycin binding (FRB) domain of FKBP12–rapamycin associated protein (FRAP) simultaneously, and the inhibition of FRAP is responsible for rapamycin's biological activity. The conformation of rapamycin in the ternary complex is very similar to that observed in the FKBP12–rapamycin binary complex, with an r.m.s. difference of only 0.30 Å. However, a slight (9°) rotation repositions the FRB-binding face of rapamycin in the ternary complex. There are extensive rapamycin–protein interactions and relatively few interactions between the two protein partners FKBP12 and FRB, these interactions mainly involving residues in the 40s and 80s loops of FKBP12 and α1 and α4 of FRB. The high-resolution refinement has revealed the crucial role of several buried waters in the formation of the ternary complex.

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Substrate processing in intramembrane proteolysis by γ-secretase – the role of protein dynamics

Biological Chemistry ◽

10.1515/hsz-2016-0269 ◽

2017 ◽

Vol 398 (4) ◽

pp. 441-453 ◽

Cited By ~ 20

Author(s):

Dieter Langosch ◽

Harald Steiner

Keyword(s):

Transmembrane Helix ◽

Peptide Bond ◽

Common Denominator ◽

Transmembrane Helices ◽

Intramembrane Proteases ◽

Bond Scission ◽

Catalytic Sites ◽

Substrate Processing

Abstract Intramembrane proteases comprise a number of different membrane proteins with different types of catalytic sites. Their common denominator is cleavage within the plane of the membrane, which usually results in peptide bond scission within the transmembrane helices of their substrates. Despite recent progress in the determination of high-resolution structures, as illustrated here for the γ-secretase complex and its substrate C99, it is still unknown how these enzymes function and how they distinguish between substrates and non-substrates. In principle, substrate/non-substrate discrimination could occur at the level of substrate binding and/or cleavage. Focusing on the γ-secretase/C99 pair, we will discuss recent observations suggesting that global motions within a substrate transmembrane helix may be much more important for defining a substrate than local unraveling at cleavage sites.

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A selective transmembrane recognition mechanism by a membrane-anchored ubiquitin ligase adaptor

The Journal of Cell Biology ◽

10.1083/jcb.202001116 ◽

2020 ◽

Vol 220 (1) ◽

Cited By ~ 1

Author(s):

Felichi Mae Arines ◽

Aaron Jeremy Hamlin ◽

Xi Yang ◽

Yun-Yu Jennifer Liu ◽

Ming Li

Keyword(s):

Conformational Changes ◽

Transmembrane Helix ◽

Environmental Cues ◽

Transmembrane Helices ◽

Recognition Mechanism ◽

Show Evidence ◽

Transmembrane Regions ◽

Underlying Mechanisms ◽

Open Question

While it is well-known that E3 ubiquitin ligases can selectively ubiquitinate membrane proteins in response to specific environmental cues, the underlying mechanisms for the selectivity are poorly understood. In particular, the role of transmembrane regions, if any, in target recognition remains an open question. Here, we describe how Ssh4, a yeast E3 ligase adaptor, recognizes the PQ-loop lysine transporter Ypq1 only after lysine starvation. We show evidence of an interaction between two transmembrane helices of Ypq1 (TM5 and TM7) and the single transmembrane helix of Ssh4. This interaction is regulated by the conserved PQ motif. Strikingly, recent structural studies of the PQ-loop family have suggested that TM5 and TM7 undergo major conformational changes during substrate transport, implying that transport-associated conformational changes may determine the selectivity. These findings thus provide critical information concerning the regulatory mechanism through which transmembrane domains can be specifically recognized in response to changing environmental conditions.

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Domains for protein-protein interactions at the N and C termini of the large subunit of bacteriophage lambda terminase.

Genetics ◽

10.1093/genetics/119.3.477 ◽

1988 ◽

Vol 119 (3) ◽

pp. 477-484

Author(s):

W F Wu ◽

S Christiansen ◽

M Feiss

Keyword(s):

Protein Interactions ◽

Ternary Complex ◽

Large Subunit ◽

Small Subunit ◽

Functional Domain ◽

Binary Complex ◽

Protein Protein Interactions ◽

Related Phage ◽

Carboxy Terminal ◽

Lambda Dna

Abstract The large subunit of phage lambda terminase, gpA, the gene product of the phage A gene, interacts with the small subunit, gpNul, to form functional terminase. Terminase binds to lambda DNA at cosB to form a binary complex. The terminase:DNA complex binds a prohead to form a ternary complex. Ternary complex formation involves an interaction of the prohead with gpA. The amino terminus of gpA contains a functional domain for interaction with gpNul, and the carboxy-terminal 38 amino acids of gpA contain a functional domain for prohead binding. This information about the structure of gpA was obtained through the use of hybrid phages resulting from recombination between lambda and the related phage 21. lambda and 21 encode terminases that are analogous in structural organization and have ca. 60% sequence identity. In spite of these similarities, lambda and 21 terminases differ in specificity for DNA binding, subunit assembly, and prohead binding. A lambda-21 hybrid phage produces a terminase in which one of the subunits is chimeric and had recombinant specificities. In the work reported here; a new hybrid, lambda-21 hybrid 67, is characterized. lambda-21 hybrid 67 is the result of a crossover between lambda and 21 in the large subunit genes, such that the DNA from the left chromosome end is from 21, including cosB phi 21, the 1 gene, and the first 48 codons for the 2 gene. The rest of the hybrid 67 chromosome is lambda DNA, including 593 codons of the A gene. The chimeric gp2/A of hybrid 67 binds gp1 to form functional terminase.(ABSTRACT TRUNCATED AT 250 WORDS)

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The Role of Selected Wavelengths of Light in the Activity of Photosystem II in Gloeobacter violaceus

International Journal of Molecular Sciences ◽

10.3390/ijms22084021 ◽

2021 ◽

Vol 22 (8) ◽

pp. 4021

Author(s):

Monika Kula-Maximenko ◽

Kamil Jan Zieliński ◽

Ireneusz Ślesak

Keyword(s):

Photosystem Ii ◽

Spectral Composition ◽

Photosynthetic Electron Transport ◽

Metabolic Energy ◽

Light Conditions ◽

Gloeobacter Violaceus ◽

Transport Chain ◽

Cyanobacterial Culture ◽

Photosynthetic Antennas

Gloeobacter violaceus is a cyanobacteria species with a lack of thylakoids, while photosynthetic antennas, i.e., phycobilisomes (PBSs), photosystem II (PSII), and I (PSI), are located in the cytoplasmic membrane. We verified the hypothesis that blue–red (BR) light supplemented with a far-red (FR), ultraviolet A (UVA), and green (G) light can affect the photosynthetic electron transport chain in PSII and explain the differences in the growth of the G. violaceus culture. The cyanobacteria were cultured under different light conditions. The largest increase in G. violaceus biomass was observed only under BR + FR and BR + G light. Moreover, the shape of the G. violaceus cells was modified by the spectrum with the addition of G light. Furthermore, it was found that both the spectral composition of light and age of the cyanobacterial culture affect the different content of phycobiliproteins in the photosynthetic antennas (PBS). Most likely, in cells grown under light conditions with the addition of FR and G light, the average antenna size increased due to the inactivation of some reaction centers in PSII. Moreover, the role of PSI and gloeorhodopsin as supplementary sources of metabolic energy in the G. violaceus growth is discussed.

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Structure and function of mitochondrial carriers - Role of the transmembrane helix P and G residues in the gating and transport mechanism

FEBS Letters ◽

10.1016/j.febslet.2009.10.063 ◽

2009 ◽

Vol 584 (9) ◽

pp. 1931-1939 ◽

Cited By ~ 75

Author(s):

Ferdinando Palmieri ◽

Ciro Leonardo Pierri

Keyword(s):

Transport Mechanism ◽

Transmembrane Helix ◽

Structure And Function ◽

Mitochondrial Carriers ◽

And Function

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Point Mutations in Transmembrane Helices 2 and 3 of ExbB and TolQ Affect Their Activities in Escherichia coli K-12

Journal of Bacteriology ◽

10.1128/jb.186.13.4402-4406.2004 ◽

2004 ◽

Vol 186 (13) ◽

pp. 4402-4406 ◽

Cited By ~ 15

Author(s):

Volkmar Braun ◽

Christina Herrmann

Keyword(s):

Escherichia Coli ◽

Amino Acid ◽

Transmembrane Helix ◽

Point Mutations ◽

Transmembrane Helices ◽

The Third ◽

Form Part ◽

K 12

ABSTRACT Replacement of glutamate 176, the only charged amino acid in the third transmembrane helix of ExbB, with alanine (E176A) abolished ExbB activity in all determined ExbB-dependent functions of Escherichia coli. Combination of the mutations T148A in the second transmembrane helix and T181A in the third transmembrane helix, proposed to form part of a proton pathway through ExbB, also resulted in inactive ExbB. E176 and T148 are strictly conserved in ExbB and TolQ proteins, and T181 is almost strictly conserved in ExbB, TolQ, and MotA.

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FOLDING ELASTIC TRANSMEMBRANE HELICES TO FIT IN A LOW-RESOLUTION IMAGE BY ELECTRON MICROSCOPY

Journal of Bioinformatics and Computational Biology ◽

10.1142/s0219720011005720 ◽

2011 ◽

Vol 09 (supp01) ◽

pp. 37-50 ◽

Cited By ~ 3

Author(s):

YUTAKA UENO ◽

KAZUNORI KAWASAKI ◽

OSAMU SAITO ◽

MASAFUMI ARAI ◽

MAKIKO SUWA

Keyword(s):

Membrane Proteins ◽

Structure Prediction ◽

Molecular Model ◽

Imaging Techniques ◽

Transmembrane Helix ◽

Prediction Method ◽

Molecular Shape ◽

Coarse Grained ◽

Transmembrane Helices ◽

Low Resolution

Structure prediction of membrane proteins could be constrained and thereby improved by introducing data of the observed molecular shape. We studied a coarse-grained molecular model that relied on residue-based dummy atoms to fold the transmembrane helices of a protein in the observed molecular shape. Based on the inter-residue potential, the α-helices were folded to contact each other in a simulated annealing protocol to search optimized conformation. Fitting the model into a three-dimensional volume was tested for proteins with known structures and resulted in a fairly reasonable arrangement of helices. In addition, the constraint to the packing transmembrane helix with the two-dimensional region was tested and found to work as a very similar folding guide. The obtained models nicely represented α-helices with the desired slight bend. Our structure prediction method for membrane proteins well demonstrated reasonable folding results using a low-resolution structural constraint introduced from recent cell-surface imaging techniques.

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TAMI-37. STEAROYL-COA DESATURASE 1 IS ESSENTIAL FOR THE GROWTH OF IDH MUTANT GLIOMA

Neuro-Oncology ◽

10.1093/neuonc/noab196.821 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi206-vi206

Author(s):

Tomohiro Yamasaki ◽

Lumin Zhang ◽

Tyrone Dowdy ◽

Adrian Lita ◽

Mark Gilbert ◽

...

Keyword(s):

Glioma Cell ◽

De Novo ◽

Glioma Cells ◽

Model Systems ◽

De Novo Lipogenesis ◽

Wild Type ◽

Knock Out ◽

Stearoyl Coa Desaturase ◽

Scd1 Expression

Abstract BACKGROUND Increased de novo lipogenesis is a hallmark of cancer metabolism. In this study, we interrogated the role of de novo lipogenesis in IDH1 mutated glioma’s growth and identified the key enzyme, Stearoyl-CoA desaturase 1 (SCD1) that provides this growth advantage. MATERIALS ANDMETHODS We prepared genetically engineered glioma cell lines (U251 wild-type: U251WT and U251 IDHR132H mutant: U251RH) and normal human astrocytes (empty vector induced-NHA: NHAEV and IDHR132H mutant: NHARH). Lipid metabolic analysis was conducted by using LC-MS and Raman imaging microscopy. SCD1 expression was investigated by The Cancer Genome Atlas (TCGA) data analysis and Western-blotting method. Knock-out of SCD1 was conducted by using CRISPR/Cas9 and shRNA. RESULTS Previously, we showed that IDH1 mut glioma cells have increased monounsaturated fatty acids (MUFAs). TCGA data revealed IDH mut glioma shows significantly higher SCD1 mRNA expression than wild-type glioma. Our model systems of IDH1 mut (U251RH, NHARH) showed increased expression of this enzyme compared with their wild-type counterpart. Moreover, addition of D-2HG to U251WT increased SCD1 expression. Herein, we showed that inhibition of SCD1 with CAY10566 decreased relative cell number and sphere forming capacity in a dose-dependent manner. Furthermore, addition of MUFAs were able to rescue the SCD1 inhibitor induced-cell death and sphere forming capacity. Knock out of SCD1 revealed decreased cell proliferation and sphere forming ability. Decreasing lipid content from the media did not alter the growth of these cells, suggesting that glioma cells rely on de novo lipid synthesis rather than scavenging them from the microenvironment. CONCLUSION Overexpression of IDH mutant gene altered lipid composition in U251 cells to enrich MUFA levels and we confirmed that D-2HG caused SCD1 upregulation in U251WT. We demonstrated the glioma cell growth requires SCD1 expression and the results of the present study may provide novel insights into the role of SCD1 in IDH mut gliomas growth.

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