National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
GM076661
United States
Other private
Research Excellence Program
United States
Citation
Journal: Elife / Year: 2019 Title: The phage L capsid decoration protein has a novel OB-fold and an unusual capsid binding strategy. Authors: Rebecca L Newcomer / Jason R Schrad / Eddie B Gilcrease / Sherwood R Casjens / Michael Feig / Carolyn M Teschke / Andrei T Alexandrescu / Kristin N Parent / Abstract: The major coat proteins of dsDNA tailed phages (order ) and herpesviruses form capsids by a mechanism that includes active packaging of the dsDNA genome into a precursor procapsid, followed by ...The major coat proteins of dsDNA tailed phages (order ) and herpesviruses form capsids by a mechanism that includes active packaging of the dsDNA genome into a precursor procapsid, followed by expansion and stabilization of the capsid. These viruses have evolved diverse strategies to fortify their capsids, such as non-covalent binding of auxiliary 'decoration' (Dec) proteins. The Dec protein from the P22-like phage L has a highly unusual binding strategy that distinguishes between nearly identical three-fold and quasi-three-fold sites of the icosahedral capsid. Cryo-electron microscopy and three-dimensional image reconstruction were employed to determine the structure of native phage L particles. NMR was used to determine the structure/dynamics of Dec in solution. The NMR structure and the cryo-EM density envelope were combined to build a model of the capsid-bound Dec trimer. Key regions that modulate the binding interface were verified by site-directed mutagenesis.
Evidence: native gel electrophoresis, We have shown that we can disrupt Dec's trimeric state by unfolding it at pH 2 for a half hour, before refolding it at pH 4, creating a monomer amenable to NMR
Type
Name
Symmetry operation
Number
identity operation
1_555
1
Buried area
0 Å2
ΔGint
0 kcal/mol
Surface area
9640 Å2
NMR ensembles
Data
Criteria
Number of conformers (submitted / calculated)
20 / 250
structures with the least restraint violations
Representative
Model #1
closest to the average
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Components
#1: Protein
Decprotein
Mass: 14374.679 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Enterobacteria phage L (virus) / Gene: dec / Plasmid: pet15b / Production host: Escherichia coli BL21 (bacteria) / Strain (production host): BL21 / References: UniProt: Q5C838
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Experimental details
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Experiment
Experiment
Method: SOLUTION NMR
NMR experiment
Conditions-ID
Experiment-ID
Solution-ID
Sample state
Spectrometer-ID
Type
1
1
3
isotropic
1
3D 1H-15N NOESY
1
2
2
isotropic
3
lrHNCO
1
3
1
isotropic
2
3D 1H-13C NOESY BB
1
4
2
isotropic
3
3D HN(CA)CB
1
5
2
isotropic
3
3D HNCO
1
6
2
isotropic
3
3D HNCA
1
7
2
isotropic
3
3DHN(CO)CA
1
8
2
isotropic
3
3DHN(CA)CO
1
9
4
isotropic
3
2D HSQC
1
10
5
isotropic
3
3D (H)CCH-TOCSY
1
11
5
isotropic
3
3D CCH-TOCSY
1
12
5
isotropic
3
3D 1H-15N TOCSY
-
Sample preparation
Details
Type
Solution-ID
Contents
Details
Label
Solvent system
solution
1
0.5 mM [U-13C; U-15N] Decoration Protein, 20 mM [U-99% 2H] sodium acetate, 50 mM sodium chloride, 100% D2O