Influenza: indentifying structural characteristics in egg -adapted vaccine preparations

The cystoviruses’ replicative mechanism and their multishell structure make them analogous to members of the reoviridae. The three dsRNA segments are termed the s, m, l segments for the small, medium and large segments respectively. They all infect strains of the plant pathogen Pseudomonas phaseolicola and are envelope by a phospholipid membrane. The receptor binding proteins, the P3 complex, attach to either host cell pili or the rough lipopolysaccharide (R-LPS) dependent on the viral species. The polymerase complex (PX) in each of the viruses is composed of four proteins, P1, P2, P4, and P7 arranged in a dodecahedral assembly that packages and then replicates the viral RNA. This sequential packaging (s, m and then the l segment) results in PX expansion accompanied by the ordered display of specific RNA receptor sites.

Our current studies utilized the techniques of cryoelecton microscopy and tomography to visualize the overall phi12 virus particle architecture. The particle was seen to have a discreet shell composed of protein P8 that defines the nucleocapsid beneath the envelope. The images suggest connection of the shell to the inner surface of the envelope. Particle reconstruction shows the P3 R-LPS attachment proteins form a circular structure termed the “donut”. We also utilized Nuclear Magnetic Resonance (NMR) spectroscopy to demonstrate that one of the PX proteins, P7, contains a flexible carboxyl terminus. This observation supports the idea that it acts as a molecular hinge facilitating complex expansion during viral RNA packaging.

Collaboration with Paul Gottlieb and Alex Wei (CUNY)

Schematic visualization of the cystovirus structure. The four proteins that form the PX are surrounded by a shell of protein P8 to form the nucleocapsid (NC). P12 is a nonstructural protein that mediates membrane acquisition by the viral particle. The murein peptidase, P5, is loosely associated with the NC. The Cryo-EM tomography data indicates that the phi12 P3 attachment proteins (P3a and P3c) form a ring-like or “donut” conformation.

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Influenza: indentifying structural characteristics in egg -adapted vaccine preparations

Schematic visualization of the cystovirus structure. The four proteins that form the PX are surrounded by a shell of protein P8 to form the nucleocapsid (NC). P12 is a nonstructural protein that mediates membrane acquisition by the viral particle. The murein peptidase, P5, is loosely associated with the NC. The Cryo-EM tomography data indicates that the phi12 P3 attachment proteins (P3a and P3c) form a ring-like or “donut” conformation.

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Influenza: indentifying structural characteristics in egg -adapted vaccine preparations

The cystoviruses, phi6 to phi14, (family cystoviridae) are a unique group of viruses that have been proven to be of great utility in the study of mechanisms extant in the field of molecular virology. With only three double stranded RNA (dsRNA) genomic segments, they all have a very similar overall genetic organization and express similar proteins. However, the amino acid sequences in the comparable proteins differ considerably among the species and thus they constitute a ready-made mutant library. The cystoviruses’ replicative mechanism and their multishell structure make them analogous to members of the reoviridae. The three dsRNA segments are termed the s, m, l segments for the small, medium and large segments respectively. They all infect strains of the plant pathogen Pseudomonas phaseolicola and are envelope by a phospholipid membrane. The receptor binding proteins, the P3 complex, attach to either host cell pili or the rough lipopolysaccharide (R-LPS) dependent on the viral species. The polymerase complex (PX) in each of the viruses is composed of four proteins, P1, P2, P4, and P7 arranged in a dodecahedral assembly that packages and then replicates the viral RNA. This sequential packaging (s, m and then the l segment) results in PX expansion accompanied by the ordered display of specific RNA receptor sites. Our current studies utilized the techniques of cryoelecton microscopy and tomography to visualize the overall phi12 virus particle architecture. The particle was seen to have a discreet shell composed of protein P8 that defines the nucleocapsid beneath the envelope. The images suggest connection of the shell to the inner surface of the envelope. Particle reconstruction shows the P3 R-LPS attachment proteins form a circular structure termed the “donut”. We also utilized Nuclear Magnetic Resonance (NMR) spectroscopy to demonstrate that one of the PX proteins, P7, contains a flexible carboxyl terminus. This observation supports the idea that it acts as a molecular hinge facilitating complex expansion during viral RNA packaging.

Collaboration with Paul Gottlieb and Alex Wei (CUNY)

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Influenza: indentifying structural characteristics in egg -adapted vaccine preparations

The cystoviruses, phi6 to phi14, (family cystoviridae) are a unique group of viruses that have been proven to be of great utility in the study of mechanisms extant in the field of molecular virology. With only three double stranded RNA (dsRNA) genomic segments, they all have a very similar overall genetic organization and express similar proteins. However, the amino acid sequences in the comparable proteins differ considerably among the species and thus they constitute a ready-made mutant library. The cystoviruses’ replicative mechanism and their multishell structure make them analogous to members of the reoviridae. The three dsRNA segments are termed the s, m, l segments for the small, medium and large segments respectively. They all infect strains of the plant pathogen Pseudomonas phaseolicola and are envelope by a phospholipid membrane. The receptor binding proteins, the P3 complex, attach to either host cell pili or the rough lipopolysaccharide (R-LPS) dependent on the viral species. The polymerase complex (PX) in each of the viruses is composed of four proteins, P1, P2, P4, and P7 arranged in a dodecahedral assembly that packages and then replicates the viral RNA. This sequential packaging (s, m and then the l segment) results in PX expansion accompanied by the ordered display of specific RNA receptor sites. Our current studies utilized the techniques of cryoelecton microscopy and tomography to visualize the overall phi12 virus particle architecture. The particle was seen to have a discreet shell composed of protein P8 that defines the nucleocapsid beneath the envelope. The images suggest connection of the shell to the inner surface of the envelope. Particle reconstruction shows the P3 R-LPS attachment proteins form a circular structure termed the “donut”. We also utilized Nuclear Magnetic Resonance (NMR) spectroscopy to demonstrate that one of the PX proteins, P7, contains a flexible carboxyl terminus. This observation supports the idea that it acts as a molecular hinge facilitating complex expansion during viral RNA packaging.

Collaboration with Paul Gottlieb and Alex Wei (CUNY)

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Influenza: indentifying structural characteristics in egg -adapted vaccine preparations

The cystoviruses, phi6 to phi14, (family cystoviridae) are a unique group of viruses that have been proven to be of great utility in the study of mechanisms extant in the field of molecular virology. With only three double stranded RNA (dsRNA) genomic segments, they all have a very similar overall genetic organization and express similar proteins. However, the amino acid sequences in the comparable proteins differ considerably among the species and thus they constitute a ready-made mutant library. The cystoviruses’ replicative mechanism and their multishell structure make them analogous to members of the reoviridae. The three dsRNA segments are termed the s, m, l segments for the small, medium and large segments respectively. They all infect strains of the plant pathogen Pseudomonas phaseolicola and are envelope by a phospholipid membrane. The receptor binding proteins, the P3 complex, attach to either host cell pili or the rough lipopolysaccharide (R-LPS) dependent on the viral species. The polymerase complex (PX) in each of the viruses is composed of four proteins, P1, P2, P4, and P7 arranged in a dodecahedral assembly that packages and then replicates the viral RNA. This sequential packaging (s, m and then the l segment) results in PX expansion accompanied by the ordered display of specific RNA receptor sites. Our current studies utilized the techniques of cryoelecton microscopy and tomography to visualize the overall phi12 virus particle architecture. The particle was seen to have a discreet shell composed of protein P8 that defines the nucleocapsid beneath the envelope. The images suggest connection of the shell to the inner surface of the envelope. Particle reconstruction shows the P3 R-LPS attachment proteins form a circular structure termed the “donut”. We also utilized Nuclear Magnetic Resonance (NMR) spectroscopy to demonstrate that one of the PX proteins, P7, contains a flexible carboxyl terminus. This observation supports the idea that it acts as a molecular hinge facilitating complex expansion during viral RNA packaging.

Collaboration with Paul Gottlieb and Alex Wei (CUNY)

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-- EdEng - 16 Nov 2012 * phi12.jpg: