G3p
From Proteopedia
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| - | In 1998, Lubkowski et al crystallized the first two domains of g3p. | + | In 1998, Lubkowski et al crystallized the first two domains of g3p. Differences from the first paper published were mainly attributed to turns in the structure and the orientation of the C terminus. The authors made two overall observations: |
(1) <scene name='G3p/Pro_213_in_cis_conformation/1'>Cis proline near C terminal end</scene> <ref name="lubkowski"/> and (2) <scene name='G3p/Oxidized_tryptophan/1'>An oxidized tryptophan</scene> <ref name="lubkowski"/>. The cis proline identified will later turn out to be important for function of the protein. The oxidized tryptophan, however, was not found in other structures <ref name="holliger 99"> PMID:10329170 </ref>. | (1) <scene name='G3p/Pro_213_in_cis_conformation/1'>Cis proline near C terminal end</scene> <ref name="lubkowski"/> and (2) <scene name='G3p/Oxidized_tryptophan/1'>An oxidized tryptophan</scene> <ref name="lubkowski"/>. The cis proline identified will later turn out to be important for function of the protein. The oxidized tryptophan, however, was not found in other structures <ref name="holliger 99"> PMID:10329170 </ref>. | ||
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The linkers don’t have a specific purpose but appear to give the protein better flexibility providing optimal infectivity <ref name="lubkowski"/>. | The linkers don’t have a specific purpose but appear to give the protein better flexibility providing optimal infectivity <ref name="lubkowski"/>. | ||
N1 interacts with TolA protein anchoring it to bacterial cell. <ref name="lubkowski"/>( and Cabilly) see Riechmann and Holliger. The C terminal domain of TolA is the coreceptor for filamentous phage infection of E coli. Cell 90, 351-360 (1997) | N1 interacts with TolA protein anchoring it to bacterial cell. <ref name="lubkowski"/>( and Cabilly) see Riechmann and Holliger. The C terminal domain of TolA is the coreceptor for filamentous phage infection of E coli. Cell 90, 351-360 (1997) | ||
| - | N1 and N2 have 15% identity <ref name="lubkowski"/> but a “nearly identical fold”, question of sharing common origins?? Possible gene duplication | ||
D2 domain binds to F pilus (Chatellier et al) | D2 domain binds to F pilus (Chatellier et al) | ||
D3 domain “anchors” to F pilus (Chatellier et al) and is necessary for phage packaging (Holliger et al) | D3 domain “anchors” to F pilus (Chatellier et al) and is necessary for phage packaging (Holliger et al) | ||
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==Evolutionarily Related Proteins== | ==Evolutionarily Related Proteins== | ||
| - | N1 and N2 have 15% identity | + | N1 and N2 have 15% identity but a “nearly identical fold”, question of sharing common origins or a gene duplication<ref name="lubkowski"/>. |
| + | |||
| + | Used DALI to identify similar proteins with both domains, Lubkowski et al were unable to identify any evolutionarily related proteins<ref name="lubkowski"/>. When looking at individual domains, they found some similar proteins. | ||
| + | |||
| + | ===D1=== | ||
| + | They reported a correlation with homopexin [[http://dx.doi.org/10.2210/pdb1hxn/pdb (1hxn)]] Z score 1.1 (p>0.05)<ref name="lubkowski"/>. A comparison with a permuted SH3 domain [[http://dx.doi.org/10.2210/pdb1tuc/pdb (1tuc)]] was made, but sequence homology did not exist<ref name="lubkowski"/>. | ||
| + | ===D2=== | ||
| + | Lubkowski et al identified the PDZ domain of Human discs large protein (1pdr) as a potentially related protein (Z score = 2.1). This protein is smaller than g3p, and consequently two beta strands in th core of domain share no identity with D2 <ref name="lubkowski"/>.(H and R, 9032075) | ||
| - | Used DALI to identify similar proteins with both domains <ref name="lubkowski"/> did not identify any | ||
| - | When looking at each domain, they found some similar proteins | ||
| - | ===N2=== | ||
| - | PDZ domain of Human discs large protein (1pdr) Z score = 2.1) while smaller, 2 beta strands in core of domain share no identity with N2 <ref name="lubkowski"/>(H and R, 9032075) | ||
PTB domain H and R, 9032075) | PTB domain H and R, 9032075) | ||
| - | ===N1=== | ||
| - | Reported correlation with homopexin [[http://dx.doi.org/10.2210/pdb1hxn/pdb (1hxn)]] Z score 1.1 (p>0.05) <ref name="lubkowski"/> | ||
| - | Compared with permuted SH3 domain [[http://dx.doi.org/10.2210/pdb1tuc/pdb (1tuc)]], but no sequence homology <ref name="lubkowski"/> | ||
==Links to Available Structures== | ==Links to Available Structures== | ||
PDB | PDB | ||
Revision as of 05:55, 28 April 2009
Contents |
Overview
Gene 3 protein (g3p pr pIII) is a minor coat protein found on the surface of filamentous bacteriophage [1]. The protein consists of 406 amino acids divided into three domains interspaced with glycine linkers [1] [2]. Peptides or proteins can be fused to g3p and evaluated for binding or other properties.
Structural Analysis
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Five major papers will be discussed outlining the evolution of structure analysis of g3p.
Initial Observations
They used NMR spectroscopy to create a structure of the first domains of g3p. Shown is a combination of the 15 most energetically favorable states. Observations of secondary structure are below.
In 1998, Lubkowski et al crystallized the first two domains of g3p. Differences from the first paper published were mainly attributed to turns in the structure and the orientation of the C terminus. The authors made two overall observations:
(1) [1] and (2) [1]. The cis proline identified will later turn out to be important for function of the protein. The oxidized tryptophan, however, was not found in other structures [3].
D1 domain
The first structure of g3p entered into the PDB was by Holliger and Riechmann in late 1996-early 1997[4]. The D1 domain consists of mostly beta sheets. Both Holliger and Riechmann as well as Lubowski et al noted a [1] in their respective publications[4][1]. This aside, five arranged as a barrel-like motif, which participates with two other strands from second domain to make an antiparallel sheet [1]. Disulfide bonds exist between Cys 7 and Cys 36 (left handed helix) and Cys 46 and Cys 53 (right handed hook) [1].
D2 domain
This domain contains eight beta strands: six in a mixed beta sheet and two interacting with D1 antiparallel sheet (β6 and β13 [1]. The amino acids between β6 and β7 doesn’t have a specific motif but has stabilizing hydrophobic interactions with other parts of the domain [1]. Three hairpins exist in this domain: between β8 and β9, β9 and β10, and β10 and β11 (cis proline in the last hairpin) [1]. The final secondary structural element is an alpha helix that interacts with rest of the domain via hydrophobic interactions [1]. Of note, there is a cation-π interaction between His 191 and Phe 199. The C terminus of D2 has seven peptides, 3 of which are proline, 1 of which is [1].
Functional Implications
Infectivity
The linkers don’t have a specific purpose but appear to give the protein better flexibility providing optimal infectivity [1]. N1 interacts with TolA protein anchoring it to bacterial cell. [1]( and Cabilly) see Riechmann and Holliger. The C terminal domain of TolA is the coreceptor for filamentous phage infection of E coli. Cell 90, 351-360 (1997) D2 domain binds to F pilus (Chatellier et al) D3 domain “anchors” to F pilus (Chatellier et al) and is necessary for phage packaging (Holliger et al) Infection with filamentous phage does not cause host cell lysis or death (Gailus and Rasched) Without N2 domain, infectivity is very low but initial contact is via N1 domain (Lubkowski et al). The complex formed by D1 and D2 may prevent destruction of the protein from bacterial proteases, Upon binding the protein opens up and D3 can then reach the inner membrane of the bacteria. (Chatellier). Horseshoe shaped molecule with polar molecules facing toward the center [1], this central area is thought to interact with pilus…however pilus is almost completely hydrophobic?? Peptides can be fused to CT domain or to the N1 domain, neither areas are near the central area of the horseshoe [1]. Amino terminus domain is necessary for infection, but full protein does not need to exist for all five particles on the surface (Cabilly). Fusions to N terminus (no affect on infectivity), between D12 and D3 (100 fold reduction for peptide insertion, and a 1000 to 100,000 fold for noncovalently interacting peptides)(Chatellier et al)
Phage Display
Insertions can also be done between D2 and D3 (H and R, 9032075)
Evolutionarily Related Proteins
N1 and N2 have 15% identity but a “nearly identical fold”, question of sharing common origins or a gene duplication[1].
Used DALI to identify similar proteins with both domains, Lubkowski et al were unable to identify any evolutionarily related proteins[1]. When looking at individual domains, they found some similar proteins.
D1
They reported a correlation with homopexin [(1hxn)] Z score 1.1 (p>0.05)[1]. A comparison with a permuted SH3 domain [(1tuc)] was made, but sequence homology did not exist[1].
D2
Lubkowski et al identified the PDZ domain of Human discs large protein (1pdr) as a potentially related protein (Z score = 2.1). This protein is smaller than g3p, and consequently two beta strands in th core of domain share no identity with D2 [1].(H and R, 9032075)
PTB domain H and R, 9032075)
Links to Available Structures
PDB [1fgp] [1g3p] [2g3p] [1tol] [1s62]
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References
- ↑ 1.00 1.01 1.02 1.03 1.04 1.05 1.06 1.07 1.08 1.09 1.10 1.11 1.12 1.13 1.14 1.15 1.16 1.17 1.18 1.19 1.20 1.21 Lubkowski J, Hennecke F, Pluckthun A, Wlodawer A. The structural basis of phage display elucidated by the crystal structure of the N-terminal domains of g3p. Nat Struct Biol. 1998 Feb;5(2):140-7. PMID:9461080
- ↑ Cabilly S. The basic structure of filamentous phage and its use in the display of combinatorial peptide libraries. Mol Biotechnol. 1999 Sep;12(2):143-8. PMID:10596371 doi:10.1385/MB:12:2:143
- ↑ Holliger P, Riechmann L, Williams RL. Crystal structure of the two N-terminal domains of g3p from filamentous phage fd at 1.9 A: evidence for conformational lability. J Mol Biol. 1999 May 14;288(4):649-57. PMID:10329170 doi:10.1006/jmbi.1999.2720
- ↑ 4.0 4.1 4.2 Holliger P, Riechmann L. A conserved infection pathway for filamentous bacteriophages is suggested by the structure of the membrane penetration domain of the minor coat protein g3p from phage fd. Structure. 1997 Feb 15;5(2):265-75. PMID:9032075
- ↑ Hill HR, Stockley PG. Phage presentation. Mol Microbiol. 1996 May;20(4):685-92. PMID:8793867 doi:10.1111/j.1365-2958.1996.tb02508.x
