IT’S ALIVE!: THE PHENOMENON OF VIRAL BREATHING

STRUCTURAL JOURNAL CLUB

AUGUST 25, 2011

WHAT IS VIRAL BREATHING?

Breathing is when the capsid undergoes reversible conformational changes not induced by an outside source.

Example: An eternal epitope is exposed on the capsid surface periodically without the presence of receptor.

DIGESTION TIME COURSE

CRYSTAL STRUCTURE OF VP4

DIGESTION IN THE PRESENCE OF DRUG

DIGESTION WITH ANTIBODY

STABILITY OF DIGESTION

Mutant: V1188M/C1199W

INTERNAL NEUTRALIZATION SITES

VP4 VP1 N terminus

TEMPERATURE DEPENDENCE

REVERSIBILITY OF INTERNAL NEUTRALIZATION

NEUTRALIZATION BY VP4 ANTIBODY

CROSS REACTIVITY OF ANTIBODIES

CVB3 STRAIN CPE

Three strains of CVB3:

- 28 – cause complete CPE on HeLa but not RD, representative strain

- RD – DAF binding strain

- RDVa – variant of RD which emerged from carrier culture

HEMAGGLUTINATION (DAF BINDING)

LIGAND BINDING OF CVB3

CVB3/28 CVB3/RD CVB3/RDVa

HeL

a

RD

Car

rier

RD

CAR

DAF

GROWTH CURVES

HeLa RD Carrier RD

CVB3/28 CVB3/RDVa CVB3/RD

CVB3 DECAY

CVB3/28 CVB3/RD CVB3/RDVa

AMINO ACID CHANGES IN CVB3

WHAT WE HAVE DONEIncubated CVB3-RD with CAR at 4 °C, RT, and 37 °C and saw the following:

4°

37°37°

RT

WHAT WE WANT TO DO

• Repeat Carson’s decay experiment to determine what the virus is doing to loose infectivity

• Negstain of different time points• Gel of time points• Western of time points

• Eventually would like to put it on affinity grid

• Virus on lipid alone without receptor to determine whether it will undergo conformation change on lipid alone

• Receptor doesn’t bind all over the virus – only one position

NOW FOR SOMETHING COMPLETELY DIFFERENT!

WARNING: YOU SHOULD NOT TRY THE FOLLOWING AT HOME! YOU WILL NEVER BE ABLE TO PUBLISH THIS!

135S AND 135S-P1

80S-P1 COMPLEXES

P1 FAB ON PV

1 OR 8 PARTICLE RECONSTRUCTION