On the catalytic mechanism of bacteriophage HK97 capsid crosslinking

Tso, Danju; Peebles, Craig L.; Maurer, Joshua B.; Duda, Robert L.; Hendrix, Roger W.

doi:10.1016/j.virol.2017.03.011

Cited by 7 publications

(7 citation statements)

References 45 publications

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“…The interaction is further strengthened by a cage of charged residues surrounding the hydrophobic pocket. This mechanism is akin to the covalent cross-linking reaction that stabilizes the HK97 capsid (15), where the site of cross-linking between polar residues K169 and N356 is protected by a hydrophobic cage composed of the amino acids leucine, methionine, and valine (48). The CUS-3 major branch of the P22-like phages may use a similar method to stabilize capsids as P22, but how the Sf6-like branch of the P22-like phages has modified this network to produce stable capsids will require further investigation.…”

Section: Discussionmentioning

confidence: 99%

A Hydrophobic Network: Intersubunit and Intercapsomer Interactions Stabilizing the Bacteriophage P22 Capsid

Asija

Teschke

2019

J Virol

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Double-stranded DNA (dsDNA) tailed phages and herpesviruses assemble their capsids using coat proteins that have the ubiquitous HK97 fold. Though this fold is common, we do not have a thorough understanding of the different ways viruses adapt it to maintain stability in various environments. The HK97-fold E-loop, which connects adjacent subunits at the outer periphery of capsomers, has been implicated in capsid stability. Here, we show that in bacteriophage P22, residue W61 at the tip of the E-loop plays a role in stabilizing procapsids and in maturation. We hypothesize that a hydrophobic pocket is formed by residues I366 and W410 in the P domain of a neighboring subunit within a capsomer, into which W61 fits like a peg. In addition, W61 likely bridges to residues A91 and L401 in P-domain loops of an adjacent capsomer, thereby linking the entire capsid together with a network of hydrophobic interactions. There is conservation of this hydrophobic network in the distantly related P22-like phages, indicating that this structural feature is likely important for stabilizing this family of phages. Thus, our data shed light on one of the varied elegant mechanisms used in nature to consistently build stable viral genome containers through subtle adaptation of the HK97 fold. IMPORTANCE Similarities in assembly reactions and coat protein structures of the dsDNA tailed phages and herpesviruses make phages ideal models to understand capsid assembly and identify potential targets for antiviral drug discovery. The coat protein E-loops of these viruses are involved in both intra- and intercapsomer interactions. In phage P22, hydrophobic interactions peg the coat protein subunits together within a capsomer, where the E-loop hydrophobic residue W61 of one subunit packs into a pocket of hydrophobic residues I366 and W410 of the adjacent subunit. W61 also makes hydrophobic interactions with A91 and L401 of a subunit in an adjacent capsomer. We show these intra- and intercapsomer hydrophobic interactions form a network crucial to capsid stability and proper assembly.

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Section: Discussionmentioning

confidence: 99%

A Hydrophobic Network: Intersubunit and Intercapsomer Interactions Stabilizing the Bacteriophage P22 Capsid

Asija

Teschke

2019

J Virol

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“…The interaction is further strengthened by a cage of charged residues surrounding the hydrophobic pocket. This mechanism is akin to the covalent crosslinking reaction that stabilize the HK97 capsid (15), where the site of crosslinking between polar residues K169 and N356 is protected by a hydrophobic cage composed of the amino acids leucine, methionine and valine (47). The CUS-3 major branch of the P22 like phages may use a similar method to stabilize capsids as P22 but how the Sf6-like branch of the P22-like phages has modified this network to produce stable capsids will require further investigation.…”

Section: Discussionmentioning

confidence: 99%

A hydrophobic network: Intersubunit and intercapsomer interactions stabilizing the bacteriophage P22 capsid

Asija

Teschke

2019

Preprint

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19dsDNA tailed phages and herpesviruses assemble their capsids using coat proteins that 20 have the ubiquitous HK97 fold. Though this fold is common, we do not have a thorough 21 understanding of the different ways viruses adapt it to maintain stability in various environments. 22The HK97-fold E-loop, which connects adjacent subunits at the outer periphery of capsomers, 23 has been implicated in capsid stability. Here we show that in bacteriophage P22, residue W61 24 at the tip of the E-loop plays a role in stabilizing procapsids and in maturation. We hypothesize 25 that a hydrophobic pocket is formed by residues I366 and W410 in the P-domain of a 26 neighboring subunit within a capsomer, into which W61 fits like a peg. In addition, W61 likely 27 bridges to residues A91 and L401 in P-domain loops of an adjacent capsomer, thereby linking 28 the entire capsid together with a network of hydrophobic interactions. There is conservation of 29 this hydrophobic network in the distantly related P22-like phages, indicating that this structural 30 feature is likely important for stabilizing this family of phages. Thus, our data shed light on one 31 of the varied elegant mechanisms used in nature to consistently build stable viral genome 32 containers through subtle adaptation of the HK97 fold.33 34 IMPORTANCE 35Similarities in assembly reactions and coat protein structures of the dsDNA tailed 36 phages and herpesviruses make phages ideal models to understand capsid assembly and 37 identify potential targets for antiviral drug discovery. The coat protein E-loops of these viruses 38 are involved in both intra-and intercapsomer interactions. In phage P22, hydrophobic 39 interactions peg the coat protein subunits together within a capsomer, where the E-loop 40 hydrophobic residue W61 of one subunit packs into a pocket of hydrophobic residues I366 and 41 W410 of the adjacent subunit. W61 also makes hydrophobic interactions with A91 and L401 of a 42 subunit in an adjacent capsomer. We show these intra-and intercapsomer hydrophobic 43 interactions form a network crucial to capsid stability and proper assembly. 48protein assembly catalyst, scaffolding protein (1). Following the formation of the DNA-free 49 precursor capsid (procapsid), the DNA is packaged through the portal ring resulting in increases 50 in the capsid volume and stability. This step is generally termed capsid expansion or maturation 51 (2). The addition of the tail machinery marks the end of assembly (2). The three-dimensional 52 arrangement of coat proteins in dsDNA viral capsids results in robust structures that can 53 withstand high levels of internal pressure that is exerted by packaged DNA, which results in 54 repulsive forces as well as the energetic strain imposed from DNA bending (3)(4)(5). In phages 55 such as phi29 the pressure inside a capsid is known to exceed 6 MPa (6). In some phages, this 56 pressure is thought to be necessary to eject the DNA into the prokaryotic host. 57The presence of a common Hong Kong 97 fold (HK97 fold) in the coat proteins of ...

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“…The formation of the isopeptide bond is catalyzed by a glutamic acid (Glu363 in HK97) located within a hydrophobic pocket made up of three amino acids (Val163, Met339, and Leu361) 24 (Figure 1). The Bobi-like isopeptide bond (Figure 8 B) is similar to that found in HK97; it is formed between lysine 70 in the E-loop and aspartic acid 266 in an adjacent major capsid protein subunit.…”

Section: The Isopeptide Bond Catalytic Site In the Bobi-like Phages I...mentioning

confidence: 99%

“…Also, the mechanism by which the isopeptide is formed may be subtly different. The catalytic glutamic acid residue is still present in the Bobi-like phages, but they lack two of the residues known to form the hydrophobic pocket that is important for the catalysis of the bond 24 . There are no obvious analogs in the Bobi-like phages for those two residues, and these phages may create the hydrophobic pocket through other means.…”

Section: Capsid Stabilitymentioning

confidence: 99%

A structural dendrogram of the actinobacteriophage major capsid proteins provides important structural insights into the evolution of capsid stability

Freeman

Gosselin

et al. 2022

Preprint

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Many double-stranded DNA viruses, including tailed bacteriophages (phages) and herpesviruses, use the HK97-fold in their major capsid protein to make the capsomers of the icosahedral viral capsid. Following the genome packaging at near-crystalline densities, the capsid is subjected to a major expansion and stabilization step that allows it to withstand environmental stresses and internal high pressure. Several different mechanisms for stabilizing the capsid have been structurally characterized, but how these mechanisms have evolved is still not understood. Using cryo-EM structure determination, structural comparisons, phylogenetic analyses, and Alphafold predictions, we have constructed a detailed structural dendrogram describing the evolution of capsid structural stability within the actinobacteriophages. The cryo-EM reconstructions of ten capsids solved to resolutions between 2.2 and 4 Angstroms revealed that eight of them exhibit major capsid proteins that are linked by a covalent cross-linking (isopeptide bond) between subunits that was first described in the HK97 phage. Those covalent interactions ultimately lead to the formation of mutually interlinked capsomers that has been compared to the structure of chain mail. However, three of the closely related phages do not exhibit such an isopeptide bond as demonstrated by both our cryo-EM maps and the lack of the required residue. This work raises questions about the importance of previously described capsid stabilization mechanisms.

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On the catalytic mechanism of bacteriophage HK97 capsid crosslinking

Cited by 7 publications

References 45 publications

A Hydrophobic Network: Intersubunit and Intercapsomer Interactions Stabilizing the Bacteriophage P22 Capsid

A Hydrophobic Network: Intersubunit and Intercapsomer Interactions Stabilizing the Bacteriophage P22 Capsid

A hydrophobic network: Intersubunit and intercapsomer interactions stabilizing the bacteriophage P22 capsid

A structural dendrogram of the actinobacteriophage major capsid proteins provides important structural insights into the evolution of capsid stability

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