The crystal structure of a bacteriophage T4 early gene product, Spackle, was determined by native sulfur single-wavelength anomalous diffraction (SAD) phasing using synchrotron radiation and was refined to 1.52 Å resolution. The structure shows that Spackle consists of a bundle of five α-helices, forming a relatively flat disc-like overall shape. Although Spackle forms a dimer in the crystal, size-exclusion chromatography with multi-angle light scattering shows that it is monomeric in solution. Mass spectrometry confirms that purified mature Spackle lacks the amino-terminal signal peptide and contains an intramolecular disulfide bond, consistent with its proposed role in the periplasm of T4 phage-infected Escherichia coli cells. The surface electrostatic potential of Spackle shows a strikingly bipolar charge distribution, suggesting a possible mode of membrane association and inhibition of the tail lysozyme activity in T4 bacteriophage superinfection exclusion.
|Original language||English (US)|
|Number of pages||6|
|Journal||Acta Crystallographica Section D: Structural Biology|
|State||Published - Sep 1 2020|
Bibliographical noteFunding Information:
This work was supported by a grant from the US National Institutes of Health (NIGMS R35-GM118047 to HA).
This work is based upon research conducted at the North-eastern Collaborative Access Team beamlines, which are funded by the US National Institutes of Health (NIGMS P30 GM124165). The PILATUS 6M detector on the 24-ID-C beamline is funded by a NIH–ORIP HEI grant (S10 RR029205). This research used resources of the Advanced Photon Source, a US Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DE-AC02-06CH11357, and those of the Minnesota Supercomputing Institute. We thank Ewa Folta-Stogniew (Biophysics Resource of Keck Laboratory at Yale University) for conducting the SEC-MALS analysis, and Todd Markowski and LeeAnn Higgins (Center for Mass Spectrometry and Proteomics, University of Minnesota) for MALDI-TOF MS analysis. The SEC-LS/UV/RI instrumentation was supported by NIH Award 1S10RR023748-01 to Ewa Folta-Stogniew.
© 2020 International Union of Crystallography. All rights reserved.
- Bacteriophage T4
- Bipolar charge distribution
- Crystal structure
- Helical bundle fold
- Native SAD phasing
- Superinfection exclusion