Supplementary data from "Glutathione transferases are structural and functional outliers in the thioredoxin fold"

Babbitt Lab > Resources > Supplementary data from "Glutathione transferases are structural and functional outliers in the thioredoxin fold"

Glutathione transferases are structural and functional outliers in the thioredoxin fold

Atkinson, HJ, and Babbitt, PC. "Glutathione transferases are structural and functional outliers in the thioredoxin fold." 2009, in preparation.

Glutathione transferases (GSTs) are ubiquitous scavengers of toxic compounds that fall, structurally and functionally, within the thioredoxin fold suprafamily. The fundamental catalytic capability of GSTs is catalysis of the nucleophilic addition or substitution of glutathione at electrophilic centers in a wide range of xenobiotic electrophilic substrates. However, little else is known about the structural and functional relationships between different subgroups of GSTs. Through a global analysis of sequence and structural similarity, it was determined that variation in the binding of glutathione between the two major classes of cytosolic GSTs results in a different mode of coenzyme activation. Additionally, the convergent features of glutathione binding between cytosolic GSTs and mitochondrial GST kappa are described. The identification of these structural and functional themes help to illuminate some of the fundamental contributions of the thioredoxin fold to catalysis, and clarify how the thioredoxin fold can be modified to enable new functions.

Available files:

Datafiles generated in the analysis
- ID lists and sequences
- protein similarity networks
- sequence alignments
Movie illustrating the similarity of the GST kappa and tau GSH binding site
Tutorials on sequence similarity networks
1. How to view SSN files
2. Basic Cytoscape tutorial

1. Datafiles generated in the analysis

Sequence files are in Fasta format (.fa)
Alignment files are in Aligned Fasta format (.afa)
Sequence similarity networks are in XGMML format
- readable using Cytoscape (http://www.cytoscape.org) — use File: Import network
- Warning: right-click or control-click network links to save to file; some browsers will interpret the .xgmml files as displayable web pages and will hang on larger files
- See tutorials for some unofficial advice on how to use similarity network files

Fig. 1 - GST structure network
Fig. 2 - GST sequence network
Fig. 3 - GST structure-based sequence alignment
Fig. 5 - Structure alignment featuring GSH sulfur position
Fig. 6 - Mitochondrial GST kappa

Fig. 1 - GST structure network

File

Description

[tab-separated text file]
gstStrucs_16.0.tsv

Information about the 40 structures present in the networks in Fig. 1.

Column Description

ID PDB ID_Chain ID

SPFamily SwissProt classification from SIMILARITY line if exists

sp_ID associated SwissProt sequence ID

exp_method NMR or X-ray

resolution if X-ray, resolution; 0.0 if NMR

[network]
gstStrucs_16.0.xgmml [290K]

Structure similarity network shown in Fig. 1 with a FAST score cutoff of 16.0

Network contains 40 structures that are a maximum of 60% identical by chain sequence and span the cytosolic GSTs. Edges at the limiting threshold of 16.0 threshold represent alignments with a median 2.5A RMSD over 177 aligned positions, while the rest of the edges represent better alignments.

This file can be viewed in Cytoscape using File: Import network

Attributes are defined for each structure in the network

Attribute:
PDB chain
Associated SwissProt sequence info
Annotation of PDB chain sequence: PFAM Trx Clan HMMs
Description

PDB chain

ID PDB ID_Chain ID

pdb_chainID PDB ID_Chain ID

pdbID PDB ID

comment PDB structure "title"

exp_method NMR or X-ray

resolution if X-ray, resolution; 0.0 if NMR

year date of deposition in the PDB

het_name list of non-standard residues (often ligands)

chain_seq amino acid sequence of PDB chain

chain_seq_length length of 'chain_seq'

chain_seq_range seq indices corresponding to SwissProt sequence

chains list of chain IDs found in entire PDB structure

Associated SwissProt sequence info

sp_ID associated SwissProt sequence ID

chain_seq_range SwissProt sequence indices indicating coverage by chain seq

AC UniProt accession

DE UniProt definition line

species species

taxID NCBI taxonomy ID

SPFamily SwissProt classification from SIMILARITY line if exists; Fig. 1 coloring

SPSequence Full length SwissProt sequence assoc w/ sp_ID

dbsource sequences is in SwissProt (sp) or TrEMBL (tr) database

seq_length SwissProt sequence length

strucIDs colon-separated list of PDB IDs associated with this SwissProt ID in the PDB database

struc_ct number of PDB IDs associated with this SwissProt ID in the PDB database

Annotation of PDB chain sequence: PFAM Trx Clan HMMs

1stModel Best hit to PFAM Trx Clan member, if aligned better than gathering threshold

1stLEval -log10(E-value) [1stModel]

domain_len length of 'domain_seq'

domain_seq sequence region aligning to '1stModel' HMM

seq_st start index in ID:sequence for 'domain_seq'

seq_end end index

Fig. 2 - GST sequence network

File

Description

[sequences]
gsts40nr1e-12.fa [174K]

622 sequences from the GST network in Fig. 2

[network]
gsts40nr1e-12.xgmml [6M]

Sequence similarity network shown in Fig. 2 with a BLAST E-value cutoff of 1x10^-12

Network contains 622 sequences that are a maximum of 40% identical and span the cytosolic GSTs. Similarity is defined by pairwise BLAST alignments better than an E-value of 1x10^-12; edges at this threshold represent alignments with a median 27% identity over 200 residues, while the rest of the edges represent better alignments.

This file can be viewed in Cytoscape using File: Import network

Attributes are defined for each sequence in the network

Attribute:

General
Domains and PFAM Trx Clan HMMs
Structures
Sequence motifs
Taxonomic
Description

General

ID UniProt:SwissProt/TrEMBL ID

name UniProt:SwissProt/TrEMBL ID

AC UniProt accession

DE UniProt definition line

SPFamily SwissProt classification from SIMILARITY line if exists; Fig 2 coloring

dbsource sequences is in SwissProt (sp) or TrEMBL (tr) database

sequence full length sequence

seq_length sequence length

Domains and PFAM Trx Clan HMMs

1stModel Best hit to PFAM Trx Clan member, if aligned better than gathering threshold

1stLEval -log10(E-value) [1stModel]

domain_seq sequence region aligning to '1stModel' HMM

domain_len length of 'domain_seq'

seq_st start index in ID:sequence for 'domain_seq'

seq_end end index

domOrder colon-separated list of Trx domains present in sequence, ordered N to C

doE-vals HMM alignment E-vals for each of the above domains

do_num_domains number of Trx clan domains present in sequence

Structures

has_struc yes if is associated with a structure in the PDB database

strucIDs colon-separated list of PDB IDs associated with this sequence in the PDB database

struc_ct number of PDB IDs associated with this sequence in the PDB database

idsFrom60nrPdbNet List of PDB chains with >= 40% identity to this sequence in the 159-structure net from Fig 1 (ie, chain sequences are also < 60% identity to each other)

nStrucsFrom60nrPdbNet = count(idsFrom60nrPdbNet)

60nrPDB_ID PDB ID for representative of this sequence in structure-based network (Fig. 1) (ie, member of the >= 40% identity sequence cluster containing the sequence associated with that PDB structure chain -- nearest by sequence from 'idsFrom60nrPdbNet')

Sequence motifs

catType CxxC, Cxxc, cxxC, loopC_C3, other;

C0 Amino acid at Cxxc position

X1 Amino acid at cXxc position

X2 Amino acid at cxXc position

C3 Amino acid at cxxC position

CXXC 'C0'+'X1'+'X2'+'C3'

cPorR is there a Pro or Arg at the N-term of the third beta strand?

Taxonomic

species species

Note that standard hierarchy is:
superkingdom, kingdom, phylum, class, order, family, genus, species

kingToSuperking If species is associated with a taxonomic kingdom, this is the attribute value. Otherwise, superkingdom value is used

phylumToClass If species is associated with a taxonomic phylum, this is the attribute value. Otherwise, class value is used.

orderToFamily If species is associated with a taxonomic order, this is the attribute value. Otherwise, family value is used

taxID NCBI taxonomy ID

Fig. 3 - GST structure-based sequence alignment

File	Description
[sequence alignment] Fig3_17strucAlignTrxDomain.afa	Sequence alignment derived from the structural alignment of 17 GST structures (Trx domain region), as shown in Fig. 3

Fig. 5 - Structure alignment featuring GSH sulfur position

File	Description
[tab-separated text file] GST_sulfurs.tsv	List of 32 GST structures with GSH or analogue sulfur visible in the alignment in Fig. 5 (does not include Grx 3: 3GRX). Text file includes info on associated SwissProt sequence, GST subgroup, ligand, etc.
[sequence alignment] 2cz2_1tu7.afa	Structure-based sequence alignment of pi GST 1TU7 and zeta GST 2CZ2 as displayed in Fig. 5. Keys the alignment of 32 structures listed in GST_sulfurs.tsv.

Fig. 6 - Mitochondrial GST kappa

File	Description
[structure network] SwissProtPfamTrxClanOnly60nr.pdb.tsv.ids_4.5.fc.xgmml [3M]	Fig.6B — structure similarity network including 159 examples spanning the full Trx fold class, thresholded at a FAST score of 4.5. For structure attributes included in the network, see network from Fig. 1 above.
[sequences] 1. Kappa-like: A.ids.fa 2. DsbA-like: B.ids.fa 3. S/C GST-like: J.ids.fa	Fig.6C — sequences used to tabulate catalytic motifs at the "CxxC" position.

2. Movie illustrating the similarity of the GST kappa and tau GSH binding site

File

Description

[Quicktime format movie]
kappa_tau_dsbG.mov [83M]
*Note: movie is pretty large; may take a while to [down]load

The movie linked at left details the similarities and differences between GST kappa (1r4w), DsbA-like DsbG (1v58), thioredoxin (1thx), and cytosolic S/C-GST tau (1oyj).

1. It begins with a view of the functional GST kappa dimer.

2. Next, thioredoxin is aligned to kappa to show where the Trx domain is located within the structure.

3. An alignment between kappa and DsbA-like DsbG is shown. While DsbG has an additional dimerization domain, there are significant similarities in the tertiary structure between kappa and DsbG; they both share an interruption at the same point within the Trx domain, and the extended "helix 2" is similar between the two enzymes. There is also sequence similarity between kappa and DsbA-like proteins within the Trx-like domain.

4. An alignment between kappa and cytosolic GST tau is shown. The overall structures are quite different.

5. However, zooming in on the bound GSH from each structure, when residues interacting with GSH are displayed, there is a surprising amount of similarity in conformation and biophysical character.

Movie created using UCSF Chimera