scholarly journals Role of Disulfide Bonds in Membrane Partitioning of a Viral Peptide

2021 ◽  
Author(s):  
Samapan Sikdar ◽  
Manidipa Banerjee ◽  
Satyavani Vemparala
Keyword(s):  
Disulfide Bond ◽  
Disulfide Bonds ◽  
Hepatitis A Virus ◽  
Host Cells ◽  
Viral Peptide ◽  
Drug Designing ◽  
Helical Hairpin ◽  
2B Protein ◽  
Compact Conformation

AbstractThe importance of disulfide bond in mediating viral peptide entry into host cells is well known. In the present work, we elucidate the role of disulfide (SS) bond in partitioning mechanism of membrane active Hepatitis A Virus-2B (HAV-2B) peptide, which harbours three cysteine residues promoting formation of multiple SS-bonded states. The inclusion of SS-bond not only results in a compact conformation but also induces distorted α-helical hairpin geometry in comparison to SS-free state, resulting in reduced hydrophobic exposure. Owing to this, the partitioning of HAV-2B peptide is completely or partly abolished. In a way, the disulfide bond regulates the partitioning of HAV-2B peptide, such that the membrane remodelling effects of this viral peptide are significantly reduced. The current findings may have potential implications in drug designing, targeting the HAV-2B protein by promoting disulfide bond formation within its membrane active region.

The Single Disulfide-Directed β-Hairpin Fold: Role of Disulfide Bond in Folding and Effect of an Additional Disulfide Bond on Stability

2020 ◽  
Vol 73 (4) ◽  
pp. 312
Author(s):  
Balasubramanyam Chittoor ◽  
Bankala Krishnarjuna ◽  
Rodrigo A. V. Morales ◽  
Raymond S. Norton
Keyword(s):  
Disulfide Bond ◽  
Disulfide Bonds ◽  
Solution Structure ◽  
Conformational Stability ◽  
Disulfide Bridge ◽  
Solution Nmr ◽  
Peptide Epitope ◽  
The Core ◽  
Native Fold

Disulfide bonds play a key role in the oxidative folding, conformational stability, and functional activity of many peptides. A few disulfide-rich peptides with privileged architecture such as the inhibitor cystine knot motif have garnered attention as templates in drug design. The single disulfide-directed β-hairpin (SDH), a novel fold identified more recently in contryphan-Vc1, has been shown to possess remarkable thermal, conformational, and chemical stability and can accept a short bioactive epitope without compromising the core structure of the peptide. In this study, we demonstrated that the single disulfide bond is critical in maintaining the native fold by replacing both cysteine residues with serine. We also designed an analogue with an additional, non-native disulfide bridge by replacing Gln1 and Tyr9 with Cys. Contryphan-Vc11–22[Q1C, Y9C] was synthesised utilising orthogonal cysteine protection and its solution structure determined using solution NMR spectroscopy. This analogue maintained the overall fold of native contryphan-Vc1. Previous studies had shown that the β-hairpin core of contryphan-Vc1 was resistant to proteolysis by trypsin and α-chymotrypsin but susceptible to cleavage by pepsin. Contryphan-Vc11–22[Q1C, Y9C] proved to be completely resistant to pepsin, thus confirming our design strategy. These results highlight the role of the disulfide bond in maintaining the SDH fold and provide a basis for the design of more stable analogues for peptide epitope grafting.

scholarly journals Effect of Cholesterol on Membrane Partitioning Dynamics of Hepatitis A Virus-2B peptide

Soft Matter ◽  
2021 ◽  
Author(s):  
Samapan Sikdar ◽  
Manidipa Banerjee ◽  
Satyavani Vemparala
Keyword(s):  
Crucial Role ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Membrane Composition ◽  
Viral Peptide ◽  
Membrane Partitioning ◽  
Peptide Detection ◽  
Host Membrane

Understanding the viral peptide detection, partitioning and subsequent host membrane composition-based response is required for gaining insights into viral mechanism. Here, we probe the crucial role of presence of membrane...

scholarly journals Redox Potentials of Disulfide Bonds in LOXL2 Studied by Nonequilibrium Alchemical Simulation

2021 ◽  
Vol 9 ◽  
Author(s):  
Lirui Lin ◽  
Haiying Zou ◽  
Wenjin Li ◽  
Li-Yan Xu ◽  
En-Min Li ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Disulfide Bond ◽  
Disulfide Bonds ◽  
Functional Role ◽  
Catalytic Domain ◽  
Lysyl Oxidase ◽  
Scavenger Receptor ◽  
Redox Potentials ◽  
Catalytic Role

Lysyl oxidase-like 2 (LOXL2) is a metalloenzyme that catalyzes the oxidative deamination ε-amino group of lysine. It is found that LOXL2 is a promotor for the metastasis and invasion of cancer cells. Disulfide bonds are important components in LOXL2, and they play a stabilizing role for protein structure or a functional role for regulating protein bioactivity. The redox potential of disulfide bond is one important property to determine the functional role of disulfide bond. In this study, we have calculated the reduction potential of all the disulfide bonds in LOXL2 by non-equilibrium alchemical simulations. Our results show that seven of seventeen disulfide bonds have high redox potentials between −182 and −298 mV and could have a functional role, viz., Cys573–Cys625, Cys579–Cys695, Cys657–Cys673, and Cys663–Cys685 in the catalytic domain, Cys351–Cys414, Cys464–Cys530, and Cys477–Cys543 in the scavenger receptor cysteine-rich (SRCR) domains. The disulfide bond of Cys351–Cys414 is predicted to play an allosteric function role, which could affect the metastasis and invasion of cancer cells. Other functional bonds have a catalytic role related to enzyme activity. The rest of disulfide bonds are predicted to play a structural role. Our study provides an important insight for the classification of disulfide bonds in LOXL2 and can be utilized for the drug design that targets the cysteine residues in LOXL2.

scholarly journals Evolution Rescues Folding of Human Immunodeficiency Virus-1 Envelope Glycoprotein GP120 Lacking a Conserved Disulfide Bond

2008 ◽  
Vol 19 (11) ◽  
pp. 4707-4716 ◽  
Author(s):  
Rogier W. Sanders ◽  
Shang-Te D. Hsu ◽  
Eelco van Anken ◽  
I. Marije Liscaljet ◽  
Martijn Dankers ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Disulfide Bonds ◽  
Side Chain ◽  
Viral Glycoprotein ◽  
Immunodeficiency Virus ◽  
Glycoprotein Gp120 ◽  
Dynamics Simulations ◽  
Β Sheet

The majority of eukaryotic secretory and membrane proteins contain disulfide bonds, which are strongly conserved within protein families because of their crucial role in folding or function. The exact role of these disulfide bonds during folding is unclear. Using virus-driven evolution we generated a viral glycoprotein variant, which is functional despite the lack of an absolutely conserved disulfide bond that links two antiparallel β-strands in a six-stranded β-barrel. Molecular dynamics simulations revealed that improved hydrogen bonding and side chain packing led to stabilization of the β-barrel fold, implying that β-sheet preference codirects glycoprotein folding in vivo. Our results show that the interactions between two β-strands that are important for the formation and/or integrity of the β-barrel can be supported by either a disulfide bond or β-sheet favoring residues.

ROLE OF DISULFIDE BONDS NEAR THE CALCIUM BINDING SITES IN FIBRINOGEN

1987 ◽  
Author(s):  
R Procyk ◽  
B Blomback
Keyword(s):  
Calcium Chloride ◽  
Disulfide Bond ◽  
Calcium Ions ◽  
Binding Sites ◽  
Calcium Binding ◽  
Disulfide Bonds ◽  
Terminal Region ◽  
Clotting Time ◽  
Calcium Binding Sites

Incubation of fibrinogen with 0.5 mM dithiothreitol in the presence of .20 mM calcium chloride cleaved disulfide bonds located at: the N-terminal end of the Aα-chain (either Aα28-Aα28 or Aα45-γ23), the C-terminal end of the Aα-chain (Aα442-Aα472) and the N-terminal end of the γ-chain (either of the symmetrical γ8, γ9 disulfides or the Aα45-γ23 disulfide bond). In the absence of calcium ions two additional disulfides, γ326-γ339, and one in the N-terminal end of the γ-chain were reduced.Plasmin digestion of the reduced fibrinogens in buffers containing calcium chloride produced fragments D and E, except that smaller fragments of D were generated from the fibrinogen in which the γ326-γ339 disulfide bonds were reduced and alkylated. In these samples calcium did not protect the C-terminal end of the γ-chain from extensive digestion.Addition of thrombin to partially reduced and alkylated fibrinogen prepared in the presence of calcium gave a clotting time similar to control unreduced fibrinogen. However, when the γ326-γ339 disulfide bonds and another γ-chain disulfide bond most likely in the N-terminal region were cleaved in reduced fibrinogen prepared in the absence of calcium, the thrombin clotting time was extremely prolonged. Apparently the disulfide bonded structure supported by γ326-γ339 was important both for binding of calcium and also for normal clotting.

scholarly journals The disulfide bond Cys2724-Cys2774 in the C-terminal cystine knot domain of von Willebrand factor is critical for its dimerization and secretion

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Yuxin Zhang ◽  
Fengwu Chen ◽  
Aizhen Yang ◽  
Xiaoying Wang ◽  
Yue Han ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Von Willebrand Factor ◽  
Disulfide Bonds ◽  
Von Willebrand Disease ◽  
Cystine Knot ◽  
Type 3 ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Background Type 3 von Willebrand disease (VWD) exhibits severe hemorrhagic tendency with complicated pathogenesis. The C-terminal cystine knot (CTCK) domain plays an important role in the dimerization and secretion of von Willebrand factor (VWF). The CTCK domain has four intrachain disulfide bonds including Cys2724-Cys2774, Cys2739-Cys2788, Cys2750-Cys2804 and Cys2754-Cys2806, and the single cysteine mutation in Cys2739-Cys2788, Cys2750-Cys2804 and Cys2754-Cys2806 result in type 3 VWD, demonstrating the crucial role of these three disulfide bonds in VWF biosynthesis, however, the role of the remaining disulfide bond Cys2724-Cys2774 remains unclear. Method and results In this study, by the next-generation sequencing we found a missense mutation a c.8171G>A (C2724Y) in the CTCK domain of VWF allele in a patient family with type 3 VWD. In vitro, VWF C2724Y protein was expressed normally in HEK-293T cells but did not form a dimer or secrete into cell culture medium, suggesting that C2724 is critical for the VWF dimerization, and thus for VWF multimerization and secretion. Conclusions Our findings provide the first genetic evidence for the important role of Cys2724-Cys2774 in VWF biosynthesis and secretion. Therefore, all of the four intrachain disulfide bonds in CTCK monomer contribute to VWF dimerization and secretion.

scholarly journals Role of Conserved Cysteines in the Alphavirus E3 Protein

2008 ◽  
Vol 83 (6) ◽  
pp. 2584-2591 ◽  
Author(s):  
Megan M. Parrott ◽  
Sarah A. Sitarski ◽  
Randy J. Arnold ◽  
Lora K. Picton ◽  
R. Blake Hill ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Viral Entry ◽  
Disulfide Bonds ◽  
Functional Role ◽  
Sindbis Virus ◽  
Virus Assembly ◽  
Wild Type ◽  
Virus Particles ◽  
Extracellular Glycoprotein

ABSTRACT Alphavirus particles are covered by 80 glycoprotein spikes that are essential for viral entry. Spikes consist of the E2 receptor binding protein and the E1 fusion protein. Spike assembly occurs in the endoplasmic reticulum, where E1 associates with pE2, a precursor containing E3 and E2 proteins. E3 is a small, cysteine-rich, extracellular glycoprotein that mediates proper folding of pE2 and its subsequent association with E1. In addition, cleavage of E3 from the assembled spike is required to make the virus particles efficiently fusion competent. We have found that the E3 protein in Sindbis virus contains one disulfide bond between residues Cys19 and Cys25. Replacing either of these two critical cysteines resulted in mutants with attenuated titers. Replacing both cysteines with either alanine or serine resulted in double mutants that were lethal. Insertion of additional cysteines based on E3 proteins from other alphaviruses resulted in either sequential or nested disulfide bond patterns. E3 sequences that formed sequential disulfides yielded virus with near-wild-type titers, while those that contained nested disulfide bonds had attenuated activity. Our data indicate that the role of the cysteine residues in E3 is not primarily structural. We hypothesize that E3 has an enzymatic or functional role in virus assembly, and these possibilities are further discussed.

The Role Of Protein Disulfide Isomerase In The Post-Ligation Phase Of β3 Integrin-Dependent Cell Adhesion

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2307-2307
Author(s):  
Avi Leader ◽  
Ronit Mor-Cohen ◽  
Nurit Rosenberg ◽  
Ron Ram ◽  
Uri Seligsohn ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Disulfide Bonds ◽  
Protein Disulfide Isomerase ◽  
Surface Expression ◽  
Inhibitory Effect ◽  
Bhk Cells ◽  
Dependent Cell ◽  
Protein Disulfide ◽  
Constitutively Active

Abstract Introduction Protein disulfide isomerase (PDI) is a member of the group of sulfhydryl isomerase enzymes which catalyze the formation, reduction and exchange of disulfide bonds. We and others have shown that PDI has a role in integrin-mediated platelet adhesion and aggregation, and among its proposed targets are integrins αIIbβ3 and αvβ3. There has been no clear evidence indicating whether disulfide bond exchange plays a role in the post-ligation phase of adhesion, which involves outside-in signaling. Objective We tested our hypothesis that free sulfhydryls and PDI play an essential role in the post-ligation phase of αIIbβ3 and αvβ3-dependent cell adhesion to fibrinogen. Methods Baby Hamster Kidney (BHK) cells were transfected with wild type (WT) αIIb and either WT β3 or β3 mutated at specific cysteines, resulting in surface expression of both αvβ3 and αIIbβ3. Single or double cysteine to serine mutations disrupting the Cys473-Cys503 and the Cys523-Cys544 disulfide bonds were generated. Evaluation of the expression and activity of WT or mutated integrins on the surface of BHK cells was performed by flow cytometry using P2 and PAC1 antibodies, respectively. Adhesion of BHK cells, expressing WT or mutated αIIbβ3, to fibrinogen-coated wells was studied in the presence or absence of bacitracin, a PDI inhibitor. Each experiment was performed with and without an αvβ3blocker (RO0655233-001). The adhered cells were stained and counted using light microscopy. Results Flow cytometry showed that BHK cells expressing the αIIbβ3 mutants bound both P2 and PAC1 while WT αIIbβ3-transfected cells bound only P2, indicating that the mutated αIIbβ3 receptors were constitutively active while WT αIIbβ3 was inactive as previously shown. The adhesion of BHK cells to fibrinogen was dependent on αIIbβ3 surface expression in both WT and mutants. Adhesion was reduced following treatment with the αvβ3 blocker, suggesting that both β3integrins support binding to immobilized fibrinogen (p < 0.05 for WT, C473S, C523S/C544S; p=0.053 for C544S). Without blocking of αvβ3, only cells expressing WT αIIbβ3 were inhibited by bacitracin (p=0.03, Figure 1A). Combination of the αvβ3 blocker and bacitracin, enabled a concentration-dependent inhibitory effect of bacitracin on adhesion of cells expressing either WT αIIbβ3or mutants disrupting the Cys523-Cys544 bond (C544S, C523S/C544S and C523S; p < 0.05 for each; Figure 1B). Statistical analysis failed to indicate a similar role for the Cys-473-Cys-503 bond (C473S and C503S; Figure 1B). Conclusion Cells expressing the constitutively active mutated αIIbβ3 were still dependent on PDI for adhesion, as shown by the inhibitory effect of the PDI inhibitor on their adhesion in the presence of an αvβ3 blocker, demonstrating that disulfide bond exchange plays an essential role in the post-ligation stage of αIIbβ3-mediated adhesion to fibrinogen. The difference between the inhibitory effect of bacitracin on the mutants disrupting the Cys523-Cys544 bond and the mutants disrupting the Cys473-Cys503 bond suggests that the role of disulfide bond exchange in the post-ligation phase of adhesion may differ between different disulfide bonds. In the absence of the αvβ3 blocker, bacitracin had no inhibitory effect on cells expressing the constitutively active mutants and the inhibitory effect of bacitracin on cells expressing the WT integrins was less prominent. Therefore, disulfide bond exchange mediated by PDI may have a pivotal role in the post-ligation phase of adhesion mediated by αIIbβ3 receptor, while the adhesion to fibrinogen mediated by αvβ3integrin depends on PDI to a lesser extent. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The Putative Herpes Simplex Virus 1 Chaperone Protein UL32 Modulates Disulfide Bond Formation during Infection

2014 ◽  
Vol 89 (1) ◽  
pp. 443-453 ◽  
Author(s):  
Brandon S. Albright ◽  
Athena Kosinski ◽  
Renata Szczepaniak ◽  
Elizabeth A. Cook ◽  
Nigel D. Stow ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Disulfide Bonds ◽  
Bond Formation ◽  
Viral Proteins ◽  
Capsid Proteins ◽  
Disulfide Bond Formation ◽  
Viral Protease ◽  
Herpes Simplex Virus 1 ◽  
Simplex Virus

ABSTRACTDuring DNA encapsidation, herpes simplex virus 1 (HSV-1) procapsids are converted to DNA-containing capsids by a process involving activation of the viral protease, expulsion of the scaffold proteins, and the uptake of viral DNA. Encapsidation requires six minor capsid proteins (UL6, UL15, UL17, UL25, UL28, and UL33) and one viral protein, UL32, not found to be associated with capsids. Although functions have been assigned to each of the minor capsid proteins, the role of UL32 in encapsidation has remained a mystery. Using an HSV-1 variant containing a functional hemagglutinin-tagged UL32, we demonstrated that UL32 was synthesized with true late kinetics and that it exhibited a previously unrecognized localization pattern. At 6 to 9 h postinfection (hpi), UL32 accumulated in viral replication compartments in the nucleus of the host cell, while at 24 hpi, it was additionally found in the cytoplasm. A newly generated UL32-null mutant was used to confirm that although B capsids containing wild-type levels of capsid proteins were synthesized, these procapsids were unable to initiate the encapsidation process. Furthermore, we showed that UL32 is redox sensitive and identified two highly conserved oxidoreductase-like C-X-X-C motifs that are essential for protein function. In addition, the disulfide bond profiles of the viral proteins UL6, UL25, and VP19C and the viral protease, VP24, were altered in the absence of UL32, suggesting that UL32 may act to modulate disulfide bond formation during procapsid assembly and maturation.IMPORTANCEAlthough functions have been assigned to six of the seven required packaging proteins of HSV, the role of UL32 in encapsidation has remained a mystery. UL32 is a cysteine-rich viral protein that contains C-X-X-C motifs reminiscent of those in proteins that participate in the regulation of disulfide bond formation. We have previously demonstrated that disulfide bonds are required for the formation and stability of the viral capsids and are also important for the formation and stability of the UL6 portal ring. In this report, we demonstrate that the disulfide bond profiles of the viral proteins UL6, UL25, and VP19C and the viral protease, VP24, are altered in cells infected with a newly isolated UL32-null mutant virus, suggesting that UL32 acts as a chaperone capable of modulating disulfide bond formation. Furthermore, these results suggest that proper regulation of disulfide bonds is essential for initiating encapsidation.

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