scholarly journals The N-Terminal Region of the Murine Coronavirus Spike Glycoprotein Is Associated with the Extended Host Range of Viruses from Persistently Infected Murine Cells

2004 ◽  
Vol 78 (17) ◽  
pp. 9073-9083 ◽  
Author(s):  
Jeanne H. Schickli ◽  
Larissa B. Thackray ◽  
Stanley G. Sawicki ◽  
Kathryn V. Holmes
Keyword(s):  
Amino Acid ◽  
Host Range ◽  
Recombinant Virus ◽  
Hepatitis Virus ◽  
Mouse Hepatitis Virus ◽  
Point Mutations ◽  
Terminal Region ◽  
Amino Acid Substitutions ◽  
Persistently Infected ◽  
Recombinant Viruses

ABSTRACT Although murine coronaviruses naturally infect only mice, several virus variants derived from persistently infected murine cell cultures have an extended host range. The mouse hepatitis virus (MHV) variant MHV/BHK can infect hamster, rat, cat, dog, monkey, and human cell lines but not the swine testis (ST) porcine cell line (J. H. Schickli, B. D. Zelus, D. E. Wentworth, S. G. Sawicki, and K. V. Holmes, J. Virol. 71:9499-9507, 1997). The spike (S) gene of MHV/BHK had 63 point mutations and a 21-bp insert that encoded 56 amino acid substitutions and a 7-amino-acid insert compared to the parental MHV strain A59. Recombinant viruses between MHV-A59 and MHV/BHK were selected in hamster cells. All of the recombinants retained 21 amino acid substitutions and a 7-amino-acid insert found in the N-terminal region of S of MHV/BHK, suggesting that these residues were responsible for the extended host range of MHV/BHK. Flow cytometry showed that MHV-A59 bound only to cells that expressed the murine glycoprotein receptor CEACAM1a. In contrast, MHV/BHK and a recombinant virus, k6c, with the 21 amino acid substitutions and 7-amino-acid insert in S bound to hamster (BHK) and ST cells as well as murine cells. Thus, 21 amino acid substitutions and a 7-amino-acid insert in the N-terminal region of the S glycoprotein of MHV/BHK confer the ability to bind and in some cases infect cells of nonmurine species.

scholarly journals Amino Acid Substitutions in the S2 Subunit of Mouse Hepatitis Virus Variant V51 Encode Determinants of Host Range Expansion

2007 ◽  
Vol 82 (3) ◽  
pp. 1414-1424 ◽  
Author(s):  
Willie C. McRoy ◽  
Ralph S. Baric
Keyword(s):  
Amino Acid ◽  
Host Range ◽  
Hepatitis Virus ◽  
Mouse Hepatitis Virus ◽  
Class I ◽  
Heptad Repeat ◽  
Amino Acid Substitutions ◽  
Spike Gene ◽  
Amino Terminal ◽  
High Affinity Receptor

ABSTRACT We previously described mouse hepatitis virus (MHV) variant V51 derived from a persistent infection of murine DBT cells with an expanded host range (R. S. Baric, E. Sullivan, L. Hensley, B. Yount, and W. Chen, J. Virol. 73:638-649, 1999). Sequencing of the V51 spike gene, the mediator of virus entry, revealed 13 amino acid substitutions relative to the originating MHV A59 strain. Seven substitutions were located in the amino-terminal S1 cleavage subunit, and six were located in the carboxy-terminal S2 cleavage subunit. Using targeted RNA recombination, we constructed a panel of recombinant viruses to map the mediators of host range to the six substitutions in S2, with a subgroup of four changes of particular interest. This subgroup maps to two previously identified domains within S2, a putative fusion peptide and a heptad repeat, both conserved features of class I fusion proteins. In addition to an altered host range, V51 displayed altered utilization of CEACAM1a, the high-affinity receptor for A59. Interestingly, a recombinant with S1 from A59 and S2 from V51 was severely debilitated in its ability to productively infect cells via CEACAM1a, while the inverse recombinant was not. This result suggests that the S2 substitutions exert powerful effects on the fusion trigger that normally passes from S1 to S2. These novel findings play against the existing data that suggest that MHV host range determinants are located in the S1 subunit, which harbors the receptor binding domain, or involve coordinating changes in both S1 and S2. Mounting evidence also suggests that the class I fusion mechanism may possess some innate plasticity that regulates viral host range.

scholarly journals Targeted Recombination within the Spike Gene of Murine Coronavirus Mouse Hepatitis Virus-A59: Q159 Is a Determinant of Hepatotropism

1998 ◽  
Vol 72 (12) ◽  
pp. 9628-9636 ◽  
Author(s):  
Isabelle Leparc-Goffart ◽  
Susan T. Hingley ◽  
Ming Ming Chua ◽  
Joanna Phillips ◽  
Ehud Lavi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Substitution ◽  
Hepatitis Virus ◽  
Mouse Hepatitis Virus ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Spike Gene ◽  
Recombinant Viruses ◽  
Directed Mutation ◽  
Murine Coronavirus

ABSTRACT Previous studies of a group of mutants of the murine coronavirus mouse hepatitis virus (MHV)-A59, isolated from persistently infected glial cells, have shown a strong correlation between a Q159L amino acid substitution in the S1 subunit of the spike gene and a loss in the ability to induce hepatitis and demyelination. To determine if Q159L alone is sufficient to cause these altered pathogenic properties, targeted RNA recombination was used to introduce a Q159L amino acid substitution into the spike gene of MHV-A59. Recombination was carried out between the genome of a temperature-sensitive mutant of MHV-A59 (Alb4) and RNA transcribed from a plasmid (pFV1) containing the spike gene as well as downstream regions, through the 3′ end, of the MHV-A59 genome. We have selected and characterized two recombinant viruses containing Q159L. These recombinant viruses (159R36 and 159R40) replicate in the brains of C57BL/6 mice and induce encephalitis to a similar extent as wild-type MHV-A59. However, they exhibit a markedly reduced ability to replicate in the liver or produce hepatitis compared to wild-type MHV-A59. These viruses also exhibit reduced virulence and reduced demyelination. A recombinant virus containing the wild-type MHV-A59 spike gene, wtR10, behaved essentially like wild-type MHV-A59. This is the first report of the isolation of recombinant viruses containing a site-directed mutation, encoding an amino acid substitution, within the spike gene of any coronavirus. This technology will allow us to begin to map the molecular determinants of pathogenesis within the spike glycoprotein.

scholarly journals Mechanism of Metronidazole Resistance inHelicobacter pylori: Comparison of the rdxA Gene Sequences in 30 Strains

2000 ◽  
Vol 44 (8) ◽  
pp. 2207-2210 ◽  
Author(s):  
Nadia Maggi Solcà ◽  
Marco Valerio Bernasconi ◽  
Jean-Claude Piffaretti
Keyword(s):  
Phylogenetic Analysis ◽  
Helicobacter Pylori ◽  
Amino Acid ◽  
Resistant Strain ◽  
Point Mutations ◽  
Housekeeping Genes ◽  
Amino Acid Substitutions ◽  
Metronidazole Resistance ◽  
Nucleotide Mutation ◽  
Nucleotide Insertion

ABSTRACT The rdxA gene of 30 independently isolatedHelicobacter pylori strains was sequenced. A comparison of the rdxA sequences revealed a higher percentage of amino acid substitutions in the corresponding protein than in other housekeeping genes. Out of 122 point mutations, 41 were missense and 4 were nonsense. A resistant strain with a nucleotide insertion in therdxA sequence was also found. With the exception of the point mutations and the insertion generating a stop signal, no particular nucleotide mutation or amino acid substitution could be associated to metronidazole resistance. Moreover, phylogenetic analysis of the 30 nucleotide sequences did not demonstrate specific clusters associated with the resistance phenotype.

scholarly journals Identification of separate domains in the adenovirus E1A gene for immortalization activity and the activation of virus early genes.

1986 ◽  
Vol 6 (10) ◽  
pp. 3470-3480 ◽  
Author(s):  
E Moran ◽  
B Zerler ◽  
T M Harrison ◽  
M B Mathews
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Point Mutations ◽  
Apparent Molecular Weight ◽  
Amino Acid Position ◽  
Cysteine Residue ◽  
Transformation Function ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
E1a Gene

The transformation and early adenovirus gene transactivation functions of the E1A region were analyzed with deletion and point mutations. Deletion of amino acids from position 86 through 120 had little effect on the lytic or transforming functions of the E1A products, while deletion of amino acids from position 121 through 150 significantly impaired both functions. The sensitivity of the transformation function to alterations in the region from amino acid position 121 to 150 was further indicated by the impairment of transforming activity resulting from single amino acid substitutions at positions 124 and 135. Interestingly, conversion of a cysteine residue at position 124 to glycine severely impaired the transformation function without affecting the early adenovirus gene activating functions. Single amino acid substitutions in a different region of the E1A gene had the converse effect. All the mutants produced polypeptides of sufficient stability to be detected by Western immunoblot analysis. The single amino acid substitutions at positions 124 and 135, although impairing the transformation functions, did not detectably alter the formation of the higher-apparent-molecular-weight forms of the E1A products.

scholarly journals Concerted Effects of Amino Acid Substitutions in Conserved Charged Residues and Other Residues in the Cytoplasmic Domain of PomA, a Stator Component of Na+-Driven Flagella

2004 ◽  
Vol 186 (20) ◽  
pp. 6749-6758 ◽  
Author(s):  
Hajime Fukuoka ◽  
Toshiharu Yakushi ◽  
Michio Homma
Keyword(s):  
Amino Acid ◽  
Temperature Shift ◽  
Terminal Region ◽  
Temperature Sensitive ◽  
Amino Acid Substitutions ◽  
Flagellar Motor ◽  
Transmembrane Segments ◽  
Suppressor Mutations ◽  
Essential Components ◽  
Charged Residues

ABSTRACT PomA is a membrane protein that is one of the essential components of the sodium-driven flagellar motor in Vibrio species. The cytoplasmic charged residues of Escherichia coli MotA, which is a PomA homolog, are believed to be required for the interaction of MotA with the C-terminal region of FliG. It was previously shown that a PomA variant with neutral substitutions in the conserved charged residues (R88A, K89A, E96Q, E97Q, and E99Q; AAQQQ) was functional. In the present study, five other conserved charged residues were replaced with neutral amino acids in the AAQQQ PomA protein. These additional substitutions did not affect the function of PomA. However, strains expressing the AAQQQ PomA variant with either an L131F or a T132M substitution, neither of which affected motor function alone, exhibited a temperature-sensitive (TS) motility phenotype. The double substitutions R88A or E96Q together with L131F were sufficient for the TS phenotype. The motility of the PomA TS mutants immediately ceased upon a temperature shift from 20 to 42°C and was restored to the original level approximately 10 min after the temperature was returned to 20°C. It is believed that PomA forms a channel complex with PomB. The complex formation of TS PomA and PomB did not seem to be affected by temperature. Suppressor mutations of the TS phenotype were mapped in the cytoplasmic boundaries of the transmembrane segments of PomA. We suggest that the cytoplasmic surface of PomA is changed by the amino acid substitutions and that the interaction of this surface with the FliG C-terminal region is temperature sensitive.

Human androgen insensitivity due to point mutations encoding amino acid substitutions in the androgen receptor steroid-binding domain

1995 ◽  
Vol 6 (2) ◽  
pp. 152-162 ◽  
Author(s):  
Koichi Murono ◽  
Berenice B. Mendonca ◽  
Ivo J. P. Arnhold ◽  
Ana C. M. M. Rigon ◽  
Claude J. Migeon ◽  
...  
Keyword(s):  
Androgen Receptor ◽  
Amino Acid ◽  
Point Mutations ◽  
Binding Domain ◽  
Amino Acid Substitutions ◽  
Androgen Insensitivity ◽  
Steroid Binding
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