Cryo-reconstructions of P22 polyheads suggest that phage assembly is nucleated by trimeric interactions among coat proteins

ABSTRACT The M13 phage assembles in the inner membrane of Escherichia coli. During maturation, about 2,700 copies of the major coat protein move from the membrane onto a single-stranded phage DNA molecule that extrudes out of the cell. The major coat protein is synthesized as a precursor, termed procoat protein, and inserts into the membrane via a Sec-independent pathway. It is processed by a leader peptidase from its leader (signal) peptide before it is assembled onto the phage DNA. The transmembrane regions of the procoat protein play an important role in all these processes. Using cysteine mutants with mutations in the transmembrane regions of the procoat and coat proteins, we investigated which of the residues are involved in multimer formation, interaction with the leader peptidase, and formation of M13 progeny particles. We found that most single cysteine residues do not interfere with the membrane insertion, processing, and assembly of the phage. Treatment of the cells with copper phenanthroline showed that the cysteine residues were readily engaged in dimer and multimer formation. This suggests that the coat proteins assemble into multimers before they proceed onto the nascent phage particles. In addition, we found that when a cysteine is located in the leader peptide at the −6 position, processing of the mutant procoat protein and of other exported proteins is affected. This inhibition of the leader peptidase results in death of the cell and shows that there are distinct amino acid residues in the M13 procoat protein involved at specific steps of the phage assembly process.

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Structure of clathrin cages and coats in ice

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010014213x ◽

1986 ◽

Vol 44 ◽

pp. 94-97

Author(s):

G.P.A. Vigers ◽

R.A. Crowther ◽

B.M.F. Pearse

Keyword(s):

Electron Microscopy ◽

Heavy Chain ◽

Outer Part ◽

Coated Vesicles ◽

Eukaryotic Cells ◽

Coat Proteins ◽

Terminal Domain ◽

Clathrin Heavy Chain ◽

Membrane Components

Clathrin forms the polyhedral cage of coated vesicles, which mediate the transfer of selected membrane components within eukaryotic cells. Clathrin cages and coated vesicles have been extensively studied by electron microscopy of negatively stained preparations and shadowed specimens. From these studies the gross morphology of the outer part of the polyhedral coat has been established and some features of the packing of clathrin trimers into the coat have also been described. However these previous studies have not revealed any internal details about the position of the terminal domain of the clathrin heavy chain, the location of the 100kd-50kd accessory coat proteins or the interactions of the coat with the enclosed membrane.

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Regulation of lipid droplet coat proteins in adipocytes in relation to insulin resistance

Endocrine Abstracts ◽

10.1530/endoabs.32.oc6.6 ◽

2013 ◽

Author(s):

Maria M Malagon ◽

Yoana Rabanal-Ruiz ◽

Rocio Guzman-Ruiz ◽

Alberto Diaz-Ruiz ◽

Andres Travez ◽

...

Keyword(s):

Insulin Resistance ◽

Lipid Droplet ◽

Coat Proteins

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Faculty Opinions recommendation of Somatic cytokinesis and pollen maturation in Arabidopsis depend on TPLATE, which has domains similar to coat proteins.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1058915.510856 ◽

2007 ◽

Author(s):

Clive Lloyd

Keyword(s):

Coat Proteins ◽

Pollen Maturation

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Faculty Opinions recommendation of Structure and organization of coat proteins in the COPII cage.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1088432.543081 ◽

2007 ◽

Author(s):

David Stephens

Keyword(s):

Coat Proteins

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Comparison of Monoclonal Gammopathies Linked to Poliovirus or Coxsackievirus vs. Other Infectious Pathogens

Cells ◽

10.3390/cells10020438 ◽

2021 ◽

Vol 10 (2) ◽

pp. 438

Author(s):

Jean Harb ◽

Nicolas Mennesson ◽

Cassandra Lepetit ◽

Maeva Fourny ◽

Margaux Louvois ◽

...

Keyword(s):

Multiple Myeloma ◽

B Cell ◽

Monoclonal Gammopathy ◽

Coat Proteins ◽

Chronic Stimulation ◽

B Cell Epitopes ◽

Monoclonal Immunoglobulins ◽

Self Antigen ◽

Coxsackievirus B1 ◽

Undetermined Significance

Chronic stimulation by infectious pathogens or self-antigen glucosylsphingosine (GlcSph) can lead to monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM). Novel assays such as the multiplex infectious antigen microarray (MIAA) and GlcSph assays, permit identification of targets for >60% purified monoclonal immunoglobulins (Igs). Searching for additional targets, we selected 28 purified monoclonal Igs whose antigen was not represented on the MIAA and GlcSph assays; their specificity of recognition was then analyzed using microarrays consisting of 3760 B-cell epitopes from 196 pathogens. The peptide sequences PALTAVETG and PALTAAETG of the VP1 coat proteins of human poliovirus 1/3 and coxsackievirus B1/B3, respectively, were specifically recognized by 6/28 monoclonal Igs. Re-analysis of patient cohorts showed that purified monoclonal Igs from 10/155 MGUS/SM (6.5%) and 3/147 MM (2.0%) bound to the PALTAVETG or PALTAAETG epitopes. Altogether, PALTAV/AETG-initiated MGUS are not rare and few seem to evolve toward myeloma.

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Biallelic variants in COPB1 cause a novel, severe intellectual disability syndrome with cataracts and variable microcephaly

Genome Medicine ◽

10.1186/s13073-021-00850-w ◽

2021 ◽

Vol 13 (1) ◽

Author(s):

William L. Macken ◽

Annie Godwin ◽

Gabrielle Wheway ◽

Karen Stals ◽

Liliya Nazlamova ◽

...

Keyword(s):

Genome Sequencing ◽

Donor Site ◽

Xenopus Tropicalis ◽

Splice Donor Site ◽

Homologous Region ◽

Disease Genes ◽

Coat Proteins ◽

Splice Donor ◽

Severe Intellectual Disability

Abstract Background Coat protein complex 1 (COPI) is integral in the sorting and retrograde trafficking of proteins and lipids from the Golgi apparatus to the endoplasmic reticulum (ER). In recent years, coat proteins have been implicated in human diseases known collectively as “coatopathies”. Methods Whole exome or genome sequencing of two families with a neuro-developmental syndrome, variable microcephaly and cataracts revealed biallelic variants in COPB1, which encodes the beta-subunit of COPI (β-COP). To investigate Family 1’s splice donor site variant, we undertook patient blood RNA studies and CRISPR/Cas9 modelling of this variant in a homologous region of the Xenopus tropicalis genome. To investigate Family 2’s missense variant, we studied cellular phenotypes of human retinal epithelium and embryonic kidney cell lines transfected with a COPB1 expression vector into which we had introduced Family 2’s mutation. Results We present a new recessive coatopathy typified by severe developmental delay and cataracts and variable microcephaly. A homozygous splice donor site variant in Family 1 results in two aberrant transcripts, one of which causes skipping of exon 8 in COPB1 pre-mRNA, and a 36 amino acid in-frame deletion, resulting in the loss of a motif at a small interaction interface between β-COP and β’-COP. Xenopus tropicalis animals with a homologous mutation, introduced by CRISPR/Cas9 genome editing, recapitulate features of the human syndrome including microcephaly and cataracts. In vitro modelling of the COPB1 c.1651T>G p.Phe551Val variant in Family 2 identifies defective Golgi to ER recycling of this mutant β-COP, with the mutant protein being retarded in the Golgi. Conclusions This adds to the growing body of evidence that COPI subunits are essential in brain development and human health and underlines the utility of exome and genome sequencing coupled with Xenopus tropicalis CRISPR/Cas modelling for the identification and characterisation of novel rare disease genes.

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Synthesis and Antiviral Evaluation of Nucleoside Analogues Bearing One Pyrimidine Moiety and Two D-Ribofuranosyl Residues

Molecules ◽

10.3390/molecules26123678 ◽

2021 ◽

Vol 26 (12) ◽

pp. 3678

Author(s):

Olga V. Andreeva ◽

Bulat F. Garifullin ◽

Vladimir V. Zarubaev ◽

Alexander V. Slita ◽

Iana L. Yesaulkova ◽

...

Keyword(s):

Influenza Virus ◽

Antiviral Activity ◽

Theoretical Calculations ◽

Nucleoside Analogs ◽

Coxsackievirus B3 ◽

Nucleoside Analogues ◽

Moderate Activity ◽

Coat Proteins ◽

Influenza Virus A ◽

Ic50 Values

A series of 1,2,3-triazolyl nucleoside analogues in which 1,2,3-triazol-4-yl-β-d-ribofuranosyl fragments are attached via polymethylene linkers to both nitrogen atoms of the heterocycle moiety (uracil, 6-methyluracil, thymine, quinazoline-2,4-dione, alloxazine) or to the C-5 and N-3 atoms of the 6-methyluracil moiety was synthesized. All compounds synthesized were evaluated for antiviral activity against influenza virus A/PR/8/34/(H1N1) and coxsackievirus B3. Antiviral assays revealed three compounds, 2i, 5i, 11c, which showed moderate activity against influenza virus A H1N1 with IC50 values of 57.5 µM, 24.3 µM, and 29.2 µM, respectively. In the first two nucleoside analogues, 1,2,3-triazol-4-yl-β-d-ribofuranosyl fragments are attached via butylene linkers to N-1 and N-3 atoms of the heterocycle moiety (6-methyluracil and alloxazine, respectively). In nucleoside analogue 11c, two 1,2,3-triazol-4-yl-2′,3′,5′-tri-O-acetyl-β-d-ribofuranose fragments are attached via propylene linkers to the C-5 and N-3 atoms of the 6-methyluracil moiety. Almost all synthesized 1,2,3-triazolyl nucleoside analogues showed no antiviral activity against the coxsackie B3 virus. Two exceptions are 1,2,3-triazolyl nucleoside analogs 2f and 5f, in which 1,2,3-triazol-4-yl-2′,3′,5′-tri-O-acetyl-β-d-ribofuranose fragments are attached to the C-5 and N-3 atoms of the heterocycle moiety (6-methyluracil and alloxazine respectively). These compounds exhibited high antiviral potency against the coxsackie B3 virus with IC50 values of 12.4 and 11.3 µM, respectively, although both were inactive against influenza virus A H1N1. According to theoretical calculations, the antiviral activity of the 1,2,3-triazolyl nucleoside analogues 2i, 5i, and 11c against the H1N1 (A/PR/8/34) influenza virus can be explained by their influence on the functioning of the polymerase acidic protein (PA) of RNA-dependent RNA polymerase (RdRp). As to the antiviral activity of nucleoside analogs 2f and 5f against coxsackievirus B3, it can be explained by their interaction with the coat proteins VP1 and VP2.

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