Sandbox Reserved 717: Difference between revisions

To finish, the phage DNA single-stranded is injected into the bacterial cytoplasm through the tail tube.

== '''Presentation of gp12''' ==

During the first step of the lytic cycle, the short tail fibers gp12 attach the virion to the host cell.  

Thanks to a three-dimensional cryoelectron microscopy, a reconstruction of the baseplate was determined to a resolution of 12 Angstrom. It has shown that the total length of the gp12 density is inferior at 340 Å. <ref name=r1>doi:10.1186/1743-422X-7-355</ref>

This technique was also a way to determine the interactions between the different proteins and their localization. Thus, interactions of the short tail fibers with each other and with the gp11 maintain the hexagon shape stability. The gp11 is also associated with gp10, which is clamped between the three fingers of gp11. This association between gp10  

Short tail fibres consist of the single protein <scene name='Sandbox_Reserved_717/Myscene/1'>gp12</scene> . This protein forms a parallel, in-register, homo-trimer of 527 residues per subunit <ref name=r3>PMID: 12888344</ref>. 18 monomers are on the baseplate. 1ocy is a monomer of the short tail fibres. A monomer of gp12 has a mass of 55.3 kDa. The gp12 N-terminal domain is bound to the baseplate. While the C-terminal globular domain is supposed to bind the bacterial host cell <ref name=r3>PMID: 12888344</ref>.

Gp12 can be divided into two fragments. One fragment with a mass of 33kDA and a second with a mass of 45kDa.  

The collar domain is a small globular domain. It contains six beta-strands and an alpha-helix <ref name=r3>PMID: 12888344</ref>.

For generating the 45kDa fragment the full length gp12 was co-expressed with its chaperone gp57 and purified <ref name=r3>PMID: 12888344</ref>. Like the 33kDa fragment it also starts with the amino acid Leu85. The 45kDa fragment contains the residues <scene name='Sandbox_Reserved_717/Mythirdscene/4'>397-517</scene> , which are in the 33kDa fragment internal deleted <ref name=r3>PMID: 12888344</ref> .

Gp12 is fixed with its N-terminal domain to the baseplate. So the C-terminal domain has to be involved in LPS-binding. To detect where the receptor-binding domain is, full-length gp12, 33kDa fragments and 45kDa fragments were immobilised in micro-plate wells and were allowed to bind to bacteria <ref name=r3>PMID: 12888344</ref>. The result was that the 33kDa fragment did never bind to a bacteria <ref name=r3>PMID: 12888344</ref>. The 45kDa fragment did bind. So the receptor-binding domain is absent in the 33kDa fragment but present in the 45kDa fragment. The residues which are present in the 45kDa fragment and lacking in the 33kDa fragment are the residues <scene name='Sandbox_Reserved_717/Mythirdscene/8'>397-517</scene>. They are referred to be part of the receptor-binding domain.

The receptor-binding domain can be sub-divided into head and bonnet. On the border between these two subdomains there is a metal-binding site <ref name=r3>PMID: 12888344</ref>. This site binds presumably to zinc. <scene name='Sandbox_Reserved_717/Mynewscene/3'>Two His amino acids </scene> (<scene name='Sandbox_Reserved_717/Mynewscene/1'>His445</scene> and <scene name='Sandbox_Reserved_717/Mynewscene/2'>His447</scene>) from each monomere are octahedrally coordinated around the zinc.  

The receptor-binding domain can be compared to a flower bud. This flower bud has got 12 petals which are organised in a 3-fold symmetry <ref name=r3>PMID: 12888344</ref>. At the bottom there are the residues 406-432 they form the  <scene name='Sandbox_Reserved_717/Mynewscene/4'>first petal </scene> , just above there are the residues 489-504 which form the <scene name='Sandbox_Reserved_717/Mynewscene/5'>second petal</scene> . The <scene name='Sandbox_Reserved_717/Mynewscene/6'>third petal </scene> is formed by the residues 450-470 and at the top there are the residues 470-480 and form the <scene name='Sandbox_Reserved_717/Mynewscene/7'>fourth petal</scene> . The complete and active receptor-binding domain is built by the trimeric protein. This trimeric proteine structure is stabilised by many inter-domain hydrogen bounds <ref name=r3>PMID: 12888344</ref>. These hydrogen bounds can have the profiles: main-chain-main-chain, main-chain-side-chain and side-chain-side-chain.

The exact position of the lipo-polysaccharide(LPS)-binding site is not known. Proteolysis experiments showed, that it contains to the domain with the residues <scene name='Sandbox_Reserved_717/Mythirdscene/8'>397-517</scene>. Compared with the homologous bacteriophage T4-like strain AR1 sequence, which also binds to the same LPS core molecule like gp12, there can be some possible binding residues be assumed.  

'''It can not be ruled out that also other amino acids are important for binding ! <ref name=r3>PMID: 12888344</ref> Further it can not be ruled out that not all of these named amino acids are important for binding.'''

The mentioned amino acids are starting point for site-directed mutations and /or other biochemical experiments to clarify the location of this binding site.

=='''Ligands and their Binding-Sites'''==

*[http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/pdbsum/GetPage.pl?pdbcode=1ocy&template=ligands.html&o=METAL&l=1.1 Click here to see the interaction between the zinc ion and the two His amino acids]

== '''Applications''' ==