title: Large-scale determination of sequence, structure, and function relationships in cytosolic glutathione transferases across the biosphere.
abstract: The cytosolic glutathione transferase (cytGST) superfamily comprises more than 13,000 nonredundant sequences found throughout the biosphere. Their key roles in metabolism and defense against oxidative damage have led to thousands of studies over several decades. Despite this attention, little is known about the physiological reactions they catalyze and most of the substrates used to assay cytGSTs are synthetic compounds. A deeper understanding of relationships across the superfamily could provide new clues about their functions. To establish a foundation for expanded classification of cytGSTs, we generated similarity-based subgroupings for the entire superfamily. Using the resulting sequence similarity networks, we chose targets that broadly covered unknown functions and report here experimental results confirming GST-like activity for 82 of them, along with 37 new 3D structures determined for 27 targets. These new data, along with experimentally known GST reactions and structures reported in the literature, were painted onto the networks to generate a global view of their sequence-structure-function relationships. The results show how proteins of both known and unknown function relate to each other across the entire superfamily and reveal that the great majority of cytGSTs have not been experimentally characterized or annotated by canonical class. A mapping of taxonomic classes across the superfamily indicates that many taxa are represented in each subgroup and highlights challenges for classification of superfamily sequences into functionally relevant classes. Experimental determination of disulfide bond reductase activity in many diverse subgroups illustrate a theme common for many reaction types. Finally, sequence comparison between an enzyme that catalyzes a reductive dechlorination reaction relevant to bioremediation efforts with some of its closest homologs reveals differences among them likely to be associated with evolution of this unusual reaction. Interactive versions of the networks, associated with functional and other types of information, can be downloaded from the Structure-Function Linkage Database (SFLD; http://sfld.rbvi.ucsf.edu).
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, California, United States of America.
authors.forename: Susan T
authors.initials: ST
authors.lastname: Mashiyama
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, United States of America.
authors.forename: M Merced
authors.initials: MM
authors.lastname: Malabanan
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, California, United States of America.
authors.forename: Eyal
authors.initials: E
authors.lastname: Akiva
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Rahul
authors.initials: R
authors.lastname: Bhosle
authors.affiliation: Department of Biochemistry, University of Wisconsin, Madison, Wisconsin, United States of America.
authors.forename: Megan C
authors.initials: MC
authors.lastname: Branch
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Brandan
authors.initials: B
authors.lastname: Hillerich
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, United States of America.
authors.forename: Kevin
authors.initials: K
authors.lastname: Jagessar
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Jungwook
authors.initials: J
authors.lastname: Kim
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Yury
authors.initials: Y
authors.lastname: Patskovsky
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Ronald D
authors.initials: RD
authors.lastname: Seidel
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Mark
authors.initials: M
authors.lastname: Stead
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Rafael
authors.initials: R
authors.lastname: Toro
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Matthew W
authors.initials: MW
authors.lastname: Vetting
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, United States of America.
authors.forename: Steven C
authors.initials: SC
authors.lastname: Almo
authors.affiliation: Departments of Biochemistry and Chemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, United States of America.
authors.forename: Richard N
authors.initials: RN
authors.lastname: Armstrong
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, California, United States of America; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, United States of America; California Institute for Quantitative Biosciences, University of California, San Francisco, San Francisco, California, United States of America.
authors.forename: Patricia C
authors.initials: PC
authors.lastname: Babbitt
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D019295:Computational Biology; D030562:Databases, Protein; D005978:Glutathione / Q000737:chemistry; D005982:Glutathione Transferase / Q000235:genetics* / Q000648:ultrastructure*; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D013329:Structure-Activity Relationship
chemical_list: D005982:Glutathione Transferase; D005978:Glutathione
references.citation: Bioinformatics. 2006 Jul 1;22(13):1658-9
references.pmid: 16731699
references.citation: Brief Bioinform. 2002 Sep;3(3):275-84
references.pmid: 12230036
references.citation: Methods Enzymol. 2005;401:169-86
references.pmid: 16399386
references.citation: Nat Genet. 2000 May;25(1):25-9
references.pmid: 10802651
references.citation: FEBS Lett. 2010 May 17;584(10):2093-101
references.pmid: 20085760
references.citation: Nucleic Acids Res. 2008 Jul 1;36(Web Server issue):W30-4
references.pmid: 18503087
references.citation: Proc Natl Acad Sci U S A. 2013 Mar 26;110(13):E1196-202
references.pmid: 23493556
references.citation: Biochemistry. 2011 Feb 22;50(7):1274-81
references.pmid: 21222452
references.citation: Curr Opin Chem Biol. 1998 Oct;2(5):618-23
references.pmid: 9818188
references.citation: Proteins. 2005 Feb 1;58(2):376-88
references.pmid: 15558583
references.citation: Annu Rev Pharmacol Toxicol. 2005;45:51-88
references.pmid: 15822171
references.citation: Protein Sci. 1999 Mar;8(3):689-92
references.pmid: 10091672
references.citation: Proteins. 2005 Feb 15;58(3):618-27
references.pmid: 15609341
references.citation: Nucleic Acids Res. 2012 Jan;40(Database issue):D565-70
references.pmid: 22123736
references.citation: Chem Biol. 2011 Oct 28;18(10):1252-60
references.pmid: 22035794
references.citation: Curr Opin Struct Biol. 2005 Dec;15(6):716-23
references.pmid: 16263269
references.citation: Nucleic Acids Res. 2014 Jan;42(Database issue):D521-30
references.pmid: 24271399
references.citation: CRC Crit Rev Biochem. 1988;23(3):283-337
references.pmid: 3069329
references.citation: Biochemistry. 2006 Feb 28;45(8):2545-55
references.pmid: 16489747
references.citation: Nucleic Acids Res. 2000 Jan 1;28(1):235-42
references.pmid: 10592235
references.citation: Biochemistry. 2009 Nov 24;48(46):11108-16
references.pmid: 19842715
references.citation: Biomol Eng. 2006 Sep;23(4):149-69
references.pmid: 16839810
references.citation: Biochem J. 2004 May 1;379(Pt 3):541-52
references.pmid: 14709161
references.citation: Genome Res. 2005 Feb;15(2):330-40
references.pmid: 15687296
references.citation: Methods Enzymol. 2005;401:1-8
references.pmid: 16399376
references.citation: Curr Opin Struct Biol. 2011 Jun;21(3):391-7
references.pmid: 21458983
references.citation: EMBO J. 1986 Apr;5(4):823-6
references.pmid: 3709526
references.citation: PLoS Comput Biol. 2009 Dec;5(12):e1000605
references.pmid: 20011109
references.citation: Bioinformatics. 2012 Nov 1;28(21):2845-6
references.pmid: 22962345
references.citation: Infect Immun. 2005 Oct;73(10):6903-11
references.pmid: 16177370
references.citation: Genome Res. 2003 Nov;13(11):2498-504
references.pmid: 14597658
references.citation: J Biol Chem. 2011 Feb 11;286(6):4271-9
references.pmid: 21106529
references.citation: Methods Enzymol. 2005;401:186-204
references.pmid: 16399387
references.citation: Biochem J. 2001 Nov 15;360(Pt 1):1-16
references.pmid: 11695986
references.citation: J Biol Chem. 2000 Aug 11;275(32):24798-806
references.pmid: 10783391
references.citation: Biochem J. 2010 May 27;428(3):419-27
references.pmid: 20388120
references.citation: Biochem J. 1997 Dec 15;328 ( Pt 3):929-35
references.pmid: 9396740
references.citation: J Biol Chem. 2012 Oct 19;287(43):35838-48
references.pmid: 22955277
references.citation: Nucleic Acids Res. 2003 Jan 1;31(1):365-70
references.pmid: 12520024
references.citation: Biochemistry. 2002 Jan 29;41(4):1308-14
references.pmid: 11802731
references.citation: PLoS Comput Biol. 2009 Oct;5(10):e1000541
references.pmid: 19851441
references.citation: Nucleic Acids Res. 2013 Jan;41(Database issue):D490-8
references.pmid: 23203873
references.citation: Biochemistry. 2004 Jan 20;43(2):352-61
references.pmid: 14717589
references.citation: Biochemistry. 2000 May 9;39(18):5303-11
references.pmid: 10820000
references.citation: Bioinformatics. 2009 May 1;25(9):1189-91
references.pmid: 19151095
references.citation: Nucleic Acids Res. 2012 Jan;40(Database issue):D790-6
references.pmid: 22102587
references.citation: Proteins. 2003 Dec 1;53(4):777-82
references.pmid: 14635120
references.citation: Bioinformatics. 1998;14(9):755-63
references.pmid: 9918945
references.citation: J Mol Biol. 2012 Jul 13;420(3):190-203
references.pmid: 22522127
references.citation: PLoS One. 2009;4(2):e4345
references.pmid: 19190775
references.citation: Biochem J. 2006 Sep 15;398(3):451-60
references.pmid: 16712527
references.citation: Nucleic Acids Res. 2012 Jan;40(Database issue):D290-301
references.pmid: 22127870
references.citation: Nucleic Acids Res. 2013 Jan;41(Database issue):D764-72
references.pmid: 23203881
references.citation: Biochemistry. 2000 Nov 14;39(45):13625-32
references.pmid: 11076500
references.citation: J Biol Chem. 2011 Mar 18;286(11):9162-73
references.pmid: 21177852
references.citation: Methods Enzymol. 2005;401:367-79
references.pmid: 16399398
references.citation: Nucleic Acids Res. 1997 Sep 1;25(17):3389-402
references.pmid: 9254694
references.citation: Drug Metab Rev. 2011 May;43(2):253-65
references.pmid: 21323601
references.citation: Nucleic Acids Res. 2002 Apr 1;30(7):1575-84
references.pmid: 11917018
references.citation: Nucleic Acids Res. 2014 Jan;42(Database issue):D304-9
references.pmid: 24304899
references.citation: Methods Enzymol. 1995;252:283-92
references.pmid: 7476363
references.citation: Chem Res Toxicol. 1997 Jan;10(1):2-18
references.pmid: 9074797
references.citation: J Mol Biol. 2006 Jul 28;360(5):934-45
references.pmid: 16806268
references.citation: Biochemistry. 2011 Nov 22;50(46):9950-62
references.pmid: 21999478
references.citation: Methods Mol Biol. 2008;426:561-75
references.pmid: 18542890
references.citation: J Mol Biol. 1998 Aug 7;281(1):135-47
references.pmid: 9680481
references.citation: J Biol Chem. 1997 Apr 25;272(17):11236-43
references.pmid: 9111025
references.citation: Nucleic Acids Res. 2008 Apr;36(7):2295-300
references.pmid: 18287115
references.citation: Biochemistry. 2009 Jul 21;48(28):6559-61
references.pmid: 19537707