The Minor Capsid Protein VP11 of Thermophilic Bacteriophage P23-77 Facilitates Virus Assembly by Using Lipid-Protein Interactions

Abstract

Thermus thermophilus bacteriophage P23-77 is the type member of a new virus family of icosahedral, tailless, inner-membranecontaining double-stranded DNA (dsDNA) viruses infecting thermophilic bacteria and halophilic archaea. The viruses have a unique capsid architecture consisting of two major capsid proteins assembled in various building blocks. We analyzed the function of the minor capsid protein VP11, which is the third known capsid component in bacteriophage P23-77. Our findings show that VP11 is a dynamically elongated dimer with a predominantly ␣-helical secondary structure and high thermal stability. The high proportion of basic amino acids in the protein enables electrostatic interaction with negatively charged molecules, including nucleic acid and large unilamellar lipid vesicles (LUVs). The plausible biological function of VP11 is elucidated by demonstrating the interactions of VP11 with Thermus-derived LUVs and with the major capsid proteins by means of the dynamic-lightscattering technique. In particular, the major capsid protein VP17 was able to link VP11-complexed LUVs into larger particles, whereas the other P23-77 major capsid protein, VP16, was unable to link VP11-comlexed LUVs. Our results rule out a previously suggested penton function for VP11. Instead, the electrostatic membrane association of VP11 triggers the binding of the major capsid protein VP17, thus facilitating a controlled incorporation of the two different major protein species into the assembling capsid. IMPORTANCEThe study of thermophilic viruses with inner membranes provides valuable insights into the mechanisms used for stabilization and assembly of protein-lipid systems at high temperatures. Our results reveal a novel way by which an internal membrane and outer capsid shell are linked in a virus that uses two different major protein species for capsid assembly. We show that a positive protein charge is important in order to form electrostatic interactions with the lipid surface, thereby facilitating the incorporation of other capsid proteins on the membrane surface. This implies an alternative function for basic proteins present in the virions of other lipid-containing thermophilic viruses, whose proposed role in genome packaging is based on their capability to bind DNA. The unique minor capsid protein of bacteriophage P23-77 resembles in its characteristics the scaffolding proteins of tailed phages, though it constitutes a substantial part of the mature virion. V iruses from extremely hot environments need to develop special strategies to cope with high temperature. Hence, they provide a valuable source for the detection of novel enzymes for biotechnology and industrial processes. Their relative simple structure makes them an ideal model system for studying life at high temperatures. With respect to extreme thermophilic bacteria, the virus studied best on the structural level is P23-77, a strictly lytic phage infecting Thermus thermophilus ATCC 33923 (1, 2, 3). P23-77 is a representative of the novel Sphaerolipoviri...

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“…Extremophilic phages, exemplified by P23-77 from alkaline hot spring and SSIP-1 originating from extreme salinity, may also constitute an important source of novel enzymes for biotechnological applications [10,97]. In addition, the structure of P23-77 suggested mechanisms for stabilization and the assembly of protein-lipid systems at extremely high temperatures [97,138]. In general, an understanding of the mechanistic properties of the membrane-containing virus particles provides interesting new insights into the engineering of tough composite nanomaterials that are well suited to protect fragile cargos [37].…”

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“…Another gene involved in encoding capsid proteins is the minor capsid protein gene. In 2015, Pawlowski et al studied functions of this gene in P23-77 phage 28 . The phage baseplate upper protein gene was also reported by Li et al in ϕ11 genome 29 .…”

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