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PMID:15701513

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Citation

Kanamaru, S, Ishiwata, Y, Suzuki, T, Rossmann, MG and Arisaka, F (2005) Control of bacteriophage T4 tail lysozyme activity during the infection process. J. Mol. Biol. 346:1013-20

Abstract

Bacteriophage T4 has an efficient mechanism for injecting the host Escherichiacoli cell with genomic DNA. Its gene product 5 (gp5) has a needle-like structure attached to the end of a tube through which the DNA passes on its way out of the head and into the host. The gp5 needle punctures the outer cell membrane and then digests the peptidoglycan cell wall in the periplasmic space. gp5 is normally post-translationally cleaved between residues 351 and 352. The function of this process in controlling the lysozyme activity of gp5 has now been investigated. When gp5 is over-expressed in E.coli, two mutants (S351H and S351A) showed a reduction of cleavage products and five other mutants (S351L, S351K, S351Y, S351Q, and S351T) showed no cleavage. Furthermore, in a complementation assay at 20 degrees C, the mutants that had no cleavage of gp5 produced a reduced number of plaques compared to wild-type T4. The crystal structure of the non-cleavage phenotype mutant of gp5, S351L, complexed with gene product 27, showed that the 18 residues in the vicinity of the potential cleavage site (disordered in the wild-type structure) had visible electron density. The polypeptide around the potential cleavage site is exposed, thus allowing access for an E.coli protease. The lysozyme activity is inhibited in the wild-type structure by a loop from the adjacent gp5 monomer that binds into the substrate-binding site. The same inhibition is apparent in the mutant structure, showing that the lysozyme is inhibited before gp5 is cleaved and, presumably, the lysozyme is activated only after gp5 has penetrated the outer membrane.

Links

PubMed Online version:10.1016/j.jmb.2004.12.042

Keywords

Bacteriophage T4/enzymology; Bacteriophage T4/genetics; Bacteriophage T4/physiology; Crystallography, X-Ray; Enzyme Activation; Enzyme Stability; Escherichia coli/virology; Genetic Complementation Test; Models, Molecular; Muramidase/chemistry; Muramidase/genetics; Muramidase/metabolism; Mutation/genetics; Protein Binding; Protein Structure, Quaternary; Serine/genetics; Serine/metabolism; Temperature

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

BPT4:NEEDL

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P17172

F

Seeded From UniProt

complete

BPT4:NEEDL

enables

GO:0042802: identical protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P16009

F

Seeded From UniProt

complete

BPT4:NEEDL

enables

GO:0003796: lysozyme activity

ECO:0000315: mutant phenotype evidence used in manual assertion

F

Seeded From UniProt

complete

BPT4:NEEDL

located_in

GO:0098015: virus tail

ECO:0000315: mutant phenotype evidence used in manual assertion

C

Seeded From UniProt

complete

BPT4:NEEDL

involved_in

GO:0046718: viral entry into host cell

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

BPT4:BP27

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P16009

F

Seeded From UniProt

complete

BPT4:BP27

located_in

GO:0098015: virus tail

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

DICDI:LYST1

GO:0003796: lysozyme activity

ECO:0000314:

F

Figure 3: This lysozyme turbidity assay describes how a S351L mutant gp5 lysozyme compares in activity to wildtype lysozyme. It depicts a more rapid increase in activity for mutant lysozymes but a greater amount of activity performed by the wildtype lysozyme.

complete

BPT4:BP27

part_of

GO:0098015: virus tail

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

BPT4:BP5

enables

GO:0042802: identical protein binding

ECO:0000315: mutant phenotype evidence used in manual assertion

F

Seeded From UniProt

complete

BPT4:BP5

involved_in

GO:0070207: protein homotrimerization

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

BPT4:BP5

involved_in

GO:0009405: pathogenesis

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

BPT4:BP5

part_of

GO:0098015: virus tail

ECO:0000315: mutant phenotype evidence used in manual assertion

C

Seeded From UniProt

complete

BPT4:BP5

enables

GO:0003796: lysozyme activity

ECO:0000315: mutant phenotype evidence used in manual assertion

F

Seeded From UniProt

complete


See also

References

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