Pages 1–8 1ytm Evolutionary trace report by report maker April 11, 2010

4.3.3 DSSP 7 4.3.4 HSSP 7 4.3.5 LaTex 7 4.3.6 Muscle 7 4.3.7 Pymol 7 4.4 Note about ET Viewer 7 4.5 Citing this work 7 4.6 About report maker 8 4.7 Attachments 8

1 INTRODUCTION From the original Protein Data Bank entry (PDB id 1ytm): Title: Crystal structure of phosphoenolpyruvate carboxykinase of succiniciproducens complexed with atp, oxalate, magnesium and manganese ions Compound: Mol id: 1; molecule: phosphoenolpyruvate carboxy- kinase [atp]; chain: a, b; synonym: pep carboxykinase; phosphoe- nolpyruvate carboxylase; pepck; ec: 4.1.1.49; engineered: yes Organism, scientific name: Anaerobiospirillum Succiniciprodu- cens; 1ytm contains a single unique chain 1ytmA (517 residues long) CONTENTS and its homologue 1ytmB.

1 Introduction 1 2 CHAIN 1YTMA 2.1 O09460 overview 2 Chain 1ytmA 1 2.1 O09460 overview 1 From SwissProt, id O09460, 97% identical to 1ytmA: 2.2 Multiple sequence alignment for 1ytmA 1 Description: Phosphoenolpyruvate carboxykinase [ATP] (EC 2.3 Residue ranking in 1ytmA 1 4.1.1.49) (PEP carboxykinase) (Phosphoenolpyruvate carboxylase) 2.4 Top ranking residues in 1ytmA and their position on (PEPCK). the structure 1 Organism, scientific name: Anaerobiospirillum succiniciprodu- 2.4.1 Clustering of residues at 25% coverage. 2 cens. 2.4.2 Overlap with known functional surfaces at : ; ; ; Aero- 25% coverage. 3 monadales; Succinivibrionaceae; Anaerobiospirillum. Catalytic activity: ATP + oxaloacetate = ADP + phosphoenolpyru- 3 Notes on using trace results 6 vate + CO(2). 3.1 Coverage 6 Pathway: Rate-limiting gluconeogenic enzyme. 3.2 Known substitutions 6 Subcellular location: Cytoplasmic (By similarity). 3.3 Surface 6 Similarity: Belongs to the phosphoenolpyruvate carboxykinase 3.4 Number of contacts 6 [ATP] family. 3.5 Annotation 6 About: This Swiss-Prot entry is copyright. It is produced through a 3.6 Mutation suggestions 6 collaboration between the Swiss Institute of Bioinformatics and the EMBL outstation - the European Bioinformatics Institute. There are 4 Appendix 6 no restrictions on its use as long as its content is in no way modified 4.1 File formats 6 and this statement is not removed. 4.2 Color schemes used 7 4.3 Credits 7 2.2 Multiple sequence alignment for 1ytmA 4.3.1 Alistat 7 For the chain 1ytmA, the alignment 1ytmA.msf (attached) with 396 4.3.2 CE 7 sequences was used. The alignment was downloaded from the HSSP

1 Lichtarge lab 2006 Fig. 1. Residues 2-259 in 1ytmA colored by their relative importance. (See Fig. 2. Residues 260-518 in 1ytmA colored by their relative importance. (See Appendix, Fig.9, for the coloring scheme.) Appendix, Fig.9, for the coloring scheme.)

database, and fragments shorter than 75% of the query as well as duplicate sequences were removed. It can be found in the attachment to this report, under the name of 1ytmA.msf. Its statistics, from the alistat program are the following:

Format: MSF Number of sequences: 396 Total number of residues: 198506 Smallest: 403 Largest: 517 Average length: 501.3 Alignment length: 517 Average identity: 49% Most related pair: 99% Most unrelated pair: 24% Most distant seq: 52%

Furthermore, 3% of residues show as conserved in this alignment. The alignment consists of 7% eukaryotic ( 3% fungi, 2% plan- tae), and 9% prokaryotic sequences. (Descriptions of some sequences were not readily available.) The file containing the sequence descrip- Fig. 3. Residues in 1ytmA, colored by their relative importance. Clockwise: tions can be found in the attachment, under the name 1ytmA.descr. front, back, top and bottom views.

2.3 Residue ranking in 1ytmA 2.4.1 Clustering of residues at 25% coverage. Fig. 4 shows the The 1ytmA sequence is shown in Figs. 1–2, with each residue colored top 25% of all residues, this time colored according to clusters they according to its estimated importance. The full listing of residues belong to. The clusters in Fig.4 are composed of the residues listed in 1ytmA can be found in the file called 1ytmA.ranks sorted in the in Table 1. attachment. Table 1. cluster size member 2.4 Top ranking residues in 1ytmA and their position on color residues the structure red 122 54,55,58,59,60,61,62,64,65 In the following we consider residues ranking among top 25% of 119,135,136,139,140,145,200 residues in the protein . Figure 3 shows residues in 1ytmA colored 201,202,203,205,206,209,213 by their importance: bright red and yellow indicate more conser- 214,217,224,225,226,227,229 ved/important residues (see Appendix for the coloring scheme). A continued in next column Pymol script for producing this figure can be found in the attachment.

2 Table 2. res type subst’s cvg noc/ dist (%) bb (A˚ ) 248 K K(100) 0.04 2/0 4.97 249 T T(100) 0.04 5/2 2.11 262 D D(99)N 0.06 5/2 3.76 263 D D(99)E 0.06 3/0 4.31 327 R R(99)Y 0.06 1/0 4.33 280 Y Y(98)T. 0.09 2/2 4.19 281 A A(98)I. 0.09 1/1 4.92

Table 2. The top 25% of residues in 1ytmA at the interface with magne- sium ion.(Field names: res: residue number in the PDB entry; type: amino acid type; substs: substitutions seen in the alignment; with the percentage of each type in the bracket; noc/bb: number of contacts with the ligand, with the number of contacts realized through backbone atoms given in the bracket; dist: distance of closest apporach to the ligand. )

Table 3. res type disruptive mutations 248 K (Y)(FTW)(SVCAG)(HD) Fig. 4. Residues in 1ytmA, colored according to the cluster they belong to: red, followed by blue and yellow are the largest clusters (see Appendix for 249 T (KR)(FQMWH)(NELPI)(D) the coloring scheme). Clockwise: front, back, top and bottom views. The 262 D (R)(FWH)(Y)(VCAG) corresponding Pymol script is attached. 263 D (R)(FWH)(YVCAG)(K) 327 R (D)(TEVLAPI)(SMCG)(FNYW) 280 Y (K)(M)(Q)(LPI) Table 1. continued 281 A (Y)(R)(KE)(H) cluster size member color residues Table 3. List of disruptive mutations for the top 25% of residues in 240,241,242,243,244,245,246 1ytmA, that are at the interface with magnesium ion. 247,248,249,250,251,252,254 257,259,260,261,262,263,264 Figure 5 shows residues in 1ytmA colored by their importance, at the 265,266,267,273,274,276,277 interface with 1ytmMG998. 278,279,280,281,282,286,291 Oxalic acid binding site. Table 4 lists the top 25% of residues 292,294,297,305,306,323,325 at the interface with 1ytmOXD543 (oxalic acid). The following table 327,330,331,335,348,354,357 (Table 5) suggests possible disruptive replacements for these residues 358,360,361,363,364,368,372 (see Section 3.6). 375,376,377,379,380,381,382 387,388,389,391,399,400,402 Table 4. 403,404,406,407,412,415,416 res type subst’s cvg noc/ dist 419,432,434,435,436,437,439 (%) bb (A˚ ) 441,443,449,450,490,507,511 206 K K(100) 0.04 15/0 3.33 514 244 S S(100) 0.04 13/5 2.32 205 K K(99)N 0.06 2/0 4.24 Table 1. Clusters of top ranking residues in 1ytmA. 263 D D(99)E 0.06 9/0 3.48 327 R R(99)Y 0.06 12/0 3.46 200 Y Y(99)F 0.07 5/0 3.66 2.4.2 Overlap with known functional surfaces at 25% coverage. 280 Y Y(98)T. 0.09 7/1 3.81 The name of the ligand is composed of the source PDB identifier 407 F F(99)L 0.09 9/0 3.45 and the heteroatom name used in that file. 60 R R(99).H 0.12 16/0 2.76 Magnesium ion binding site. Table 2 lists the top 25% of residues 225 H H(97) 0.14 8/0 3.46 at the interface with 1ytmMG998 (magnesium ion). The following L(1)Y table (Table 3) suggests possible disruptive replacements for these 388 T T(96) 0.18 2/1 4.35 residues (see Section 3.6). V(2)L.I continued in next column

3 Table 5. continued res type disruptive mutations

Table 5. List of disruptive mutations for the top 25% of residues in 1ytmA, that are at the interface with oxalic acid.

Fig. 5. Residues in 1ytmA, at the interface with magnesium ion, colored by their relative importance. The ligand (magnesium ion) is colored green. Atoms further than 30A˚ away from the geometric center of the ligand, as well as on the line of sight to the ligand were removed. (See Appendix for the coloring scheme for the protein chain 1ytmA.)

Table 4. continued res type subst’s cvg noc/ dist (%) bb (A˚ ) Fig. 6. Residues in 1ytmA, at the interface with oxalic acid, colored by their S relative importance. The ligand (oxalic acid) is colored green. Atoms further than 30A˚ away from the geometric center of the ligand, as well as on the line Table 4. The top 25% of residues in 1ytmA at the interface with oxalic of sight to the ligand were removed. (See Appendix for the coloring scheme acid.(Field names: res: residue number in the PDB entry; type: amino acid for the protein chain 1ytmA.) type; substs: substitutions seen in the alignment; with the percentage of each type in the bracket; noc/bb: number of contacts with the ligand, with the num- ber of contacts realized through backbone atoms given in the bracket; dist: Figure 6 shows residues in 1ytmA colored by their importance, at the distance of closest apporach to the ligand. ) interface with 1ytmOXD543. ATP binding site. Table 6 lists the top 25% of residues at the inter- face with 1ytmATP541 (atp). The following table (Table 7) suggests Table 5. possible disruptive replacements for these residues (see Section 3.6). res type disruptive mutations Table 6. 206 K (Y)(FTW)(SVCAG)(HD) res type subst’s cvg noc/ dist 244 S (KR)(FQMWH)(NYELPI)(D) (%) bb (A˚ ) 205 K (Y)(FTW)(SVCAG)(H) 206 K K(100) 0.04 1/0 4.78 263 D (R)(FWH)(YVCAG)(K) 242 G G(100) 0.04 1/1 4.89 327 R (D)(TEVLAPI)(SMCG)(FNYW) 243 L L(100) 0.04 7/7 3.69 200 Y (K)(Q)(EM)(NR) 244 S S(100) 0.04 24/18 3.39 280 Y (K)(M)(Q)(LPI) 245 G G(100) 0.04 33/33 2.55 407 F (KE)(T)(QDR)(SCG) 247 G G(100) 0.04 33/33 3.11 60 R (TD)(SEVCLAPIG)(YM)(FNW) 248 K K(100) 0.04 39/16 2.75 225 H (E)(Q)(KM)(TD) 249 T T(100) 0.04 32/17 2.89 388 T (R)(K)(H)(Q) 251 L L(100) 0.04 7/1 3.95 continued in next column 246 T T(99)S 0.06 27/25 3.38 continued in next column

4 Table 6. continued Table 7. continued res type subst’s cvg noc/ dist res type disruptive (%) bb (A˚ ) mutations 262 D D(99)N 0.06 1/1 4.72 388 T (R)(K)(H)(Q) 263 D D(99)E 0.06 6/0 2.94 446 I (R)(Y)(H)(KE) 327 R R(99)Y 0.06 16/0 2.66 280 Y Y(98)T. 0.09 1/1 4.06 Table 7. List of disruptive mutations for the top 25% of residues in 282 K K(99). 0.09 30/3 3.45 1ytmA, that are at the interface with ATP. 434 N N(99).S 0.10 1/1 4.94 435 T T(98)S. 0.10 29/16 3.62 V 250 T T(99)S 0.11 33/7 2.81 443 R R(99)TL 0.11 45/0 3.06 225 H H(97) 0.14 4/0 3.42 L(1)Y 449 T T(96) 0.16 10/0 2.97 S(3)AC 388 T T(96) 0.18 1/0 4.43 V(2)L.I S 446 I L(39) 0.25 31/6 3.48 I(59)GV

Table 6. The top 25% of residues in 1ytmA at the interface with ATP.(Field names: res: residue number in the PDB entry; type: amino acid type; substs: substitutions seen in the alignment; with the percentage of each type in the bracket; noc/bb: number of contacts with the ligand, with the num- ber of contacts realized through backbone atoms given in the bracket; dist: distance of closest apporach to the ligand. )

Table 7. res type disruptive mutations Fig. 7. Residues in 1ytmA, at the interface with ATP, colored by their relative 206 K (Y)(FTW)(SVCAG)(HD) importance. The ligand (ATP) is colored green. Atoms further than 30A˚ away 242 G (KER)(FQMWHD)(NYLPI)(SVA) from the geometric center of the ligand, as well as on the line of sight to the 243 L (YR)(TH)(SKECG)(FQWD) ligand were removed. (See Appendix for the coloring scheme for the protein 244 S (KR)(FQMWH)(NYELPI)(D) chain 1ytmA.) 245 G (KER)(FQMWHD)(NYLPI)(SVA) 247 G (KER)(FQMWHD)(NYLPI)(SVA) 248 K (Y)(FTW)(SVCAG)(HD) Figure 7 shows residues in 1ytmA colored by their importance, at the 249 T (KR)(FQMWH)(NELPI)(D) interface with 1ytmATP541. 251 L (YR)(TH)(SKECG)(FQWD) Manganese (ii) ion binding site. Table 8 lists the top 25% of resi- 246 T (KR)(FQMWH)(NELPI)(D) dues at the interface with 1ytmMN999 (manganese (ii) ion). The 262 D (R)(FWH)(Y)(VCAG) following table (Table 9) suggests possible disruptive replacements 263 D (R)(FWH)(YVCAG)(K) for these residues (see Section 3.6). 327 R (D)(TEVLAPI)(SMCG)(FNYW) Table 8. 280 Y (K)(M)(Q)(LPI) res type subst’s cvg noc/ dist 282 K (Y)(FTW)(SVCAG)(HD) (%) bb (A˚ ) 434 N (Y)(FWH)(TR)(EVCAG) 206 K K(100) 0.04 3/0 2.48 435 T (KR)(QH)(FMW)(NE) 244 S S(100) 0.04 2/1 4.58 250 T (KR)(FQMWH)(NELPI)(D) 248 K K(100) 0.04 2/0 4.07 443 R (D)(TYE)(FVCLAWPIG)(S) 263 D D(99)E 0.06 4/0 2.25 225 H (E)(Q)(KM)(TD) 407 F F(99)L 0.09 2/0 4.67 449 T (KR)(QH)(FMW)(E) 225 H H(97) 0.14 5/0 2.27 continued in next column L(1)Y continued in next column

5 Table 8. continued 3 NOTES ON USING TRACE RESULTS res type subst’s cvg noc/ dist 3.1 Coverage (%) bb (A˚ ) Trace results are commonly expressed in terms of coverage: the resi- due is important if its “coverage” is small - that is if it belongs to Table 8. The top 25% of residues in 1ytmA at the interface with manga- nese (ii) ion.(Field names: res: residue number in the PDB entry; type: amino some small top percentage of residues [100% is all of the residues acid type; substs: substitutions seen in the alignment; with the percentage of in a chain], according to trace. The ET results are presented in the each type in the bracket; noc/bb: number of contacts with the ligand, with form of a table, usually limited to top 25% percent of residues (or the number of contacts realized through backbone atoms given in the bracket; to some nearby percentage), sorted by the strength of the presumed dist: distance of closest apporach to the ligand. ) evolutionary pressure. (I.e., the smaller the coverage, the stronger the pressure on the residue.) Starting from the top of that list, mutating a couple of residues should affect the protein somehow, with the exact effects to be determined experimentally. Table 9. res type disruptive 3.2 Known substitutions mutations One of the table columns is “substitutions” - other amino acid types 206 K (Y)(FTW)(SVCAG)(HD) seen at the same position in the alignment. These amino acid types 244 S (KR)(FQMWH)(NYELPI)(D) may be interchangeable at that position in the protein, so if one wants 248 K (Y)(FTW)(SVCAG)(HD) to affect the protein by a point mutation, they should be avoided. For 263 D (R)(FWH)(YVCAG)(K) example if the substitutions are “RVK” and the original protein has 407 F (KE)(T)(QDR)(SCG) an R at that position, it is advisable to try anything, but RVK. Conver- 225 H (E)(Q)(KM)(TD) sely, when looking for substitutions which will not affect the protein, one may try replacing, R with K, or (perhaps more surprisingly), with Table 9. List of disruptive mutations for the top 25% of residues in V. The percentage of times the substitution appears in the alignment 1ytmA, that are at the interface with manganese (ii) ion. is given in the immediately following bracket. No percentage is given in the cases when it is smaller than 1%. This is meant to be a rough guide - due to rounding errors these percentages often do not add up to 100%.

3.3 Surface To detect candidates for novel functional interfaces, first we look for residues that are solvent accessible (according to DSSP program) by 2 at least 10A˚ , which is roughly the area needed for one water mole- cule to come in the contact with the residue. Furthermore, we require that these residues form a “cluster” of residues which have neighbor within 5A˚ from any of their heavy atoms. Note, however, that, if our picture of protein evolution is correct, the neighboring residues which are not surface accessible might be equally important in maintaining the interaction specificity - they should not be automatically dropped from consideration when choo- sing the set for mutagenesis. (Especially if they form a cluster with the surface residues.)

3.4 Number of contacts Another column worth noting is denoted “noc/bb”; it tells the num- ber of contacts heavy atoms of the residue in question make across the interface, as well as how many of them are realized through the backbone atoms (if all or most contacts are through the backbone, mutation presumably won’t have strong impact). Two heavy atoms are considered to be “in contact” if their centers are closer than 5A˚ . Fig. 8. Residues in 1ytmA, at the interface with manganese (ii) ion, colored by their relative importance. The ligand (manganese (ii) ion) is colored green. Atoms further than 30A˚ away from the geometric center of the ligand, as well 3.5 Annotation as on the line of sight to the ligand were removed. (See Appendix for the If the residue annotation is available (either from the pdb file or coloring scheme for the protein chain 1ytmA.) from other sources), another column, with the header “annotation” appears. Annotations carried over from PDB are the following: site (indicating existence of related site record in PDB ), S-S (disulfide Figure 8 shows residues in 1ytmA colored by their importance, at the bond forming residue), hb (hydrogen bond forming residue, jb (james interface with 1ytmMN999. bond forming residue), and sb (for salt bridge forming residue).

6 3.6 Mutation suggestions Mutation suggestions are completely heuristic and based on comple- mentarity with the substitutions found in the alignment. Note that they are meant to be disruptive to the interaction of the protein with its ligand. The attempt is made to complement the following COVERAGE properties: small [AV GSTC], medium [LPNQDEMIK], large [WFYHR], hydrophobic [LPVAMWFI], polar [GTCY ]; posi- V tively [KHR], or negatively [DE] charged, aromatic [WFYH], 100% 50% 30% 5% long aliphatic chain [EKRQM], OH-group possession [SDETY ], and NH2 group possession [NQRK]. The suggestions are listed according to how different they appear to be from the original amino acid, and they are grouped in round brackets if they appear equally disruptive. From left to right, each bracketed group of amino acid V types resembles more strongly the original (i.e. is, presumably, less disruptive) These suggestions are tentative - they might prove disrup- RELATIVE IMPORTANCE tive to the fold rather than to the interaction. Many researcher will choose, however, the straightforward alanine mutations, especially in the beginning stages of their investigation. Fig. 9. Coloring scheme used to color residues by their relative importance.

4 APPENDIX alignment length (e.g. including gap characters). Also shown are 4.1 File formats some percent identities. A percent pairwise alignment identity is defi- Files with extension “ranks sorted” are the actual trace results. The ned as (idents / MIN(len1, len2)) where idents is the number of fields in the table in this file: exact identities and len1, len2 are the unaligned lengths of the two sequences. The ”average percent identity”, ”most related pair”, and • alignment# number of the position in the alignment ”most unrelated pair” of the alignment are the average, maximum, • residue# residue number in the PDB file and minimum of all (N)(N-1)/2 pairs, respectively. The ”most distant seq” is calculated by finding the maximum pairwise identity (best • type amino acid type relative) for all N sequences, then finding the minimum of these N • rank rank of the position according to older version of ET numbers (hence, the most outlying sequence). alistat is copyrighted • variability has two subfields: by HHMI/Washington University School of Medicine, 1992-2001, 1. number of different amino acids appearing in in this column and freely distributed under the GNU General Public License. of the alignment 4.3.2 CE To map ligand binding sites from different 2. their type source structures, report maker uses the CE program: • rho ET score - the smaller this value, the lesser variability of http://cl.sdsc.edu/. Shindyalov IN, Bourne PE (1998) this position across the branches of the tree (and, presumably, ”Protein structure alignment by incremental combinatorial extension the greater the importance for the protein) (CE) of the optimal path . Protein Engineering 11(9) 739-747. • cvg coverage - percentage of the residues on the structure which 4.3.3 DSSP In this work a residue is considered solvent accessi- have this rho or smaller ble if the DSSP program finds it exposed to water by at least 10A˚ 2, • gaps percentage of gaps in this column which is roughly the area needed for one water molecule to come in the contact with the residue. DSSP is copyrighted by W. Kabsch, C. 4.2 Color schemes used Sander and MPI-MF, 1983, 1985, 1988, 1994 1995, CMBI version The following color scheme is used in figures with residues colored by [email protected] November 18,2002, by cluster size: black is a single-residue cluster; clusters composed of http://www.cmbi.kun.nl/gv/dssp/descrip.html. more than one residue colored according to this hierarchy (ordered by descending size): red, blue, yellow, green, purple, azure, tur- 4.3.4 HSSP Whenever available, report maker uses HSSP ali- quoise, brown, coral, magenta, LightSalmon, SkyBlue, violet, gold, gnment as a starting point for the analysis (sequences shorter than bisque, LightSlateBlue, orchid, RosyBrown, MediumAquamarine, 75% of the query are taken out, however); R. Schneider, A. de DarkOliveGreen, CornflowerBlue, grey55, burlywood, LimeGreen, Daruvar, and C. Sander. ”The HSSP database of protein structure- tan, DarkOrange, DeepPink, maroon, BlanchedAlmond. sequence alignments.” Nucleic Acids Res., 25:226–230, 1997. The colors used to distinguish the residues by the estimated evolutionary pressure they experience can be seen in Fig. 9. http://swift.cmbi.kun.nl/swift/hssp/ 4.3 Credits 4.3.5 LaTex The text for this report was processed using LATEX; Leslie Lamport, “LaTeX: A Document Preparation System Addison- 4.3.1 Alistat alistat reads a multiple sequence alignment from the Wesley,” Reading, Mass. (1986). file and shows a number of simple statistics about it. These stati- stics include the format, the number of sequences, the total number 4.3.6 Muscle When making alignments “from scratch”, report of residues, the average and range of the sequence lengths, and the maker uses Muscle alignment program: Edgar, Robert C. (2004),

7 ”MUSCLE: multiple sequence alignment with high accuracy and of service for comparative analysis of proteins.” Bioinformatics high throughput.” Nucleic Acids Research 32(5), 1792-97. 22:1656-7. http://www.drive5.com/muscle/ 4.6 About report maker 4.3.7 Pymol The figures in this report were produced using report maker was written in 2006 by Ivana Mihalek. The 1D ran- Pymol. The scripts can be found in the attachment. Pymol king visualization program was written by Ivica Res.ˇ report maker is an open-source application copyrighted by DeLano Scien- is copyrighted by Lichtarge Lab, Baylor College of Medicine, tific LLC (2005). For more information about Pymol see Houston. http://pymol.sourceforge.net/. (Note for Windows 4.7 Attachments users: the attached package needs to be unzipped for Pymol to read The following files should accompany this report: the scripts and launch the viewer.) • 1ytmA.complex.pdb - coordinates of 1ytmA with all of its 4.4 Note about ET Viewer interacting partners Dan Morgan from the Lichtarge lab has developed a visualization • 1ytmA.etvx - ET viewer input file for 1ytmA tool specifically for viewing trace results. If you are interested, please • visit: 1ytmA.cluster report.summary - Cluster report summary for 1ytmA http://mammoth.bcm.tmc.edu/traceview/ • 1ytmA.ranks - Ranks file in sequence order for 1ytmA The viewer is self-unpacking and self-installing. Input files to be used • 1ytmA.clusters - Cluster descriptions for 1ytmA with ETV (extension .etvx) can be found in the attachment to the • 1ytmA.msf - the multiple sequence alignment used for the chain main report. 1ytmA 4.5 Citing this work • 1ytmA.descr - description of sequences used in 1ytmA msf The method used to rank residues and make predictions in this report • 1ytmA.ranks sorted - full listing of residues and their ranking can be found in Mihalek, I., I. Res,ˇ O. Lichtarge. (2004). ”A Family of for 1ytmA Evolution-Entropy Hybrid Methods for Ranking of Protein Residues • 1ytmA.1ytmMG998.if.pml - Pymol script for Figure 5 by Importance” J. Mol. Bio. 336: 1265-82. For the original version • 1ytmA.cbcvg - used by other 1ytmA – related pymol scripts of ET see O. Lichtarge, H.Bourne and F. Cohen (1996). ”An Evolu- • tionary Trace Method Defines Binding Surfaces Common to Protein 1ytmA.1ytmOXD543.if.pml - Pymol script for Figure 6 Families” J. Mol. Bio. 257: 342-358. • 1ytmA.1ytmATP541.if.pml - Pymol script for Figure 7 report maker itself is described in Mihalek I., I. Res and O. • 1ytmA.1ytmMN999.if.pml - Pymol script for Figure 8 Lichtarge (2006). ”Evolutionary Trace Report Maker: a new type

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