Pages 1–8 2fbm Evolutionary trace report by report maker September 19, 2008

4.3.1 Alistat 7 4.3.2 CE 7 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 Data Bank entry (PDB id 2fbm): Title: Acetyltransferase domain of cdy1 Compound: Mol id: 1; molecule: y protein 1, telomeric isoform b; chain: a, b, c; engineered: yes Organism, scientific name: Homo Sapiens; 2fbm contains a single unique chain 2fbmA (251 residues long) and its homologues 2fbmC and 2fbmB. CONTENTS 2 CHAIN 2FBMA 1 Introduction 1 2.1 Q9Y6F8 overview 2 Chain 2fbmA 1 From SwissProt, id Q9Y6F8, 100% identical to 2fbmA: Description: 2.1 Q9Y6F8 overview 1 Testis-specific chromodomain protein Y 1. Organism, scientific name: 2.2 Multiple sequence alignment for 2fbmA 1 Homo sapiens (Human). Taxonomy: 2.3 Residue ranking in 2fbmA 1 Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; 2.4 Top ranking residues in 2fbmA and their position on Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Primates; the structure 2 Catarrhini; Hominidae; Homo. Subcellular location: 2.4.1 Clustering of residues at 25% coverage. 2 Nuclear (By similarity). Alternative products: 2.4.2 Overlap with known functional surfaces at 25% coverage. 2 Event=Alternative splicing; Named isoforms=2; Name=1; 2.4.3 Possible novel functional surfaces at 25% IsoId=Q9Y6F8-1; Sequence=Displayed; Name=2; IsoId=Q9Y6F8- coverage. 4 2; Sequence=VSP 001079; Tissue specificity: Testis specific. 3 Notes on using trace results 6 Similarity: Contains 1 chromo domain. 3.1 Coverage 6 About: This Swiss-Prot entry is copyright. It is produced through a 3.2 Known substitutions 6 collaboration between the Swiss Institute of Bioinformatics and the 3.3 Surface 6 EMBL outstation - the European Bioinformatics Institute. There are 3.4 Number of contacts 6 no restrictions on its use as long as its content is in no way modified 3.5 Annotation 6 and this statement is not removed. 3.6 Mutation suggestions 6 2.2 Multiple sequence alignment for 2fbmA 4 Appendix 6 For the chain 2fbmA, the alignment 2fbmA.msf (attached) with 91 4.1 File formats 6 sequences was used. The alignment was assembled through combi- 4.2 Color schemes used 7 nation of BLAST searching on the UniProt database and alignment 4.3 Credits 7 using Muscle program. It can be found in the attachment to this

1 Lichtarge lab 2006 Pymol script for producing this figure can be found in the attachment.

Fig. 1. Residues 282-406 in 2fbmA colored by their relative importance. (See Appendix, Fig.8, for the coloring scheme.)

Fig. 2. Residues 407-532 in 2fbmA colored by their relative importance. (See Appendix, Fig.8, for the coloring scheme.) report, under the name of 2fbmA.msf. Its statistics, from the alistat program are the following: Fig. 3. Residues in 2fbmA, colored by their relative importance. Clockwise: Format: MSF front, back, top and bottom views. Number of sequences: 91 Total number of residues: 22176 Smallest: 212 2.4.1 Clustering of residues at 25% coverage. Fig. 4 shows the Largest: 251 top 25% of all residues, this time colored according to clusters they Average length: 243.7 belong to. The clusters in Fig.4 are composed of the residues listed Alignment length: 251 in Table 1. Average identity: 30% Most related pair: 99% Table 1. Most unrelated pair: 16% cluster size member Most distant seq: 30% color residues red 62 297,299,300,302,303,307,308 309,327,333,335,336,341,343 Furthermore, <1% of residues show as conserved in this ali- 344,345,346,381,382,385,386 gnment. 387,388,389,391,392,393,394 The alignment consists of 36% eukaryotic ( 26% vertebrata, 1% 395,397,399,402,403,405,407 arthropoda, 4% fungi, 1% plantae), 50% prokaryotic, and 12% 411,413,416,417,421,424,425 archaean sequences. (Descriptions of some sequences were not rea- 427,428,429,432,437,441,453 dily available.) The file containing the sequence descriptions can be 455,456,460,461,462,465,475 found in the attachment, under the name 2fbmA.descr. 479,494,505,512,528,532 2.3 Residue ranking in 2fbmA Table 1. Clusters of top ranking residues in 2fbmA. The 2fbmA sequence is shown in Figs. 1–2, with each residue colored according to its estimated importance. The full listing of residues in 2fbmA can be found in the file called 2fbmA.ranks sorted in the 2.4.2 Overlap with known functional surfaces at 25% coverage. attachment. The name of the ligand is composed of the source PDB identifier 2.4 Top ranking residues in 2fbmA and their position on and the heteroatom name used in that file. Interface with 2fbmC. the structure Table 2 lists the top 25% of residues at the interface with 2fbmC. The following table (Table 3) suggests possible In the following we consider residues ranking among top 25% of disruptive replacements for these residues (see Section 3.6). residues in the protein . Figure 3 shows residues in 2fbmA colored by their importance: bright red and yellow indicate more conser- ved/important residues (see Appendix for the coloring scheme). A

2 Table 2. continued res type subst’s cvg noc/ dist (%) bb (A˚ ) 405 V I(20) 0.19 5/3 4.25 V(43) R(30) A(3) L(1) 462 V V(74) 0.23 2/2 4.00 A(14) L(3) I(5) C(2)

Table 2. The top 25% of residues in 2fbmA at the interface with 2fbmC. (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. Fig. 4. Residues in 2fbmA, colored according to the cluster they belong to: res type disruptive red, followed by blue and yellow are the largest clusters (see Appendix for the coloring scheme). Clockwise: front, back, top and bottom views. The mutations corresponding Pymol script is attached. 403 D (R)(K)(FW)(H) 382 P (Y)(R)(H)(T) 460 G (E)(D)(FW)(KYR) Table 2. 461 L (R)(Y)(T)(H) res type subst’s cvg noc/ dist 402 C (K)(ER)(Q)(D) (%) bb (A˚ ) 405 V (Y)(E)(R)(K) 403 D D(97) 0.00 39/28 2.83 462 V (R)(Y)(KE)(H) H(1) T(1) Table 3. List of disruptive mutations for the top 25% of residues in 382 P P(93) 0.03 2/0 4.27 2fbmA, that are at the interface with 2fbmC. I(3) V(1) Figure 5 shows residues in 2fbmA colored by their importance, at the L(1) interface with 2fbmC. M(1) Interface with 2fbmB.Table 4 lists the top 25% of residues at the 460 G G(95) 0.04 13/13 4.03 interface with 2fbmB. The following table (Table 5) suggests possible N(1) disruptive replacements for these residues (see Section 3.6). Q(1) R(1) Table 4. H(1) res type subst’s cvg noc/ dist 461 L L(83) 0.11 1/1 4.78 (%) bb (A˚ ) F(3) 424 P P(80) 0.05 13/3 3.75 I(9) A(14) M(1) N(2) V(2) L(2) 402 C C(71) 0.13 9/9 2.85 S(1) S(4) 437 G G(87) 0.10 8/8 3.96 G(7) S(7) F(7) N(2) A(8) V(1) continued in next column P(1) 512 E E(81) 0.10 6/3 4.19 continued in next column

3 Table 4. continued res type subst’s cvg noc/ dist (%) bb (A˚ ) H(2) C(1) F(2) I(1)

Table 4. The top 25% of residues in 2fbmA at the interface with 2fbmB. (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 5. res type disruptive mutations 424 P (YR)(H)(T)(KE) 437 G (R)(KE)(H)(FYW) 512 E (H)(Y)(FW)(R) 417 Y (K)(Q)(E)(R) Fig. 5. Residues in 2fbmA, at the interface with 2fbmC, colored by their rela- tive importance. 2fbmC is shown in backbone representation (See Appendix 421 G (E)(K)(R)(D) for the coloring scheme for the protein chain 2fbmA.) 429 S (KR)(Q)(E)(H)

Table 5. List of disruptive mutations for the top 25% of residues in Table 4. continued 2fbmA, that are at the interface with 2fbmB. res type subst’s cvg noc/ dist (%) bb (A˚ ) A(3) M(5) .(3) P(1) G(1) L(1) V(1) S(1) R(1) 417 Y E(42) 0.14 1/1 4.73 F(37) Y(9) W(2) H(3) L(1) C(1) N(1) A(1) 421 G G(87) 0.14 7/7 3.77 A(6) R(3) H(1) S(1) 429 S S(40) 0.20 28/13 3.69 Fig. 6. Residues in 2fbmA, at the interface with 2fbmB, colored by their rela- T(40) tive importance. 2fbmB is shown in backbone representation (See Appendix A(2) for the coloring scheme for the protein chain 2fbmA.) L(9) continued in next column Figure 6 shows residues in 2fbmA colored by their importance, at the interface with 2fbmB.

4 2.4.3 Possible novel functional surfaces at 25% coverage. One Table 6. continued group of residues is conserved on the 2fbmA surface, away from (or res type substitutions(%) cvg susbtantially larger than) other functional sites and interfaces reco- .(3)P(1)G(1) gnizable in PDB entry 2fbm. It is shown in Fig. 7. The right panel L(1)V(1)S(1) shows (in blue) the rest of the larger cluster this surface belongs to. R(1) 461 L L(83)F(3)I(9) 0.11 M(1)V(2) 302 R R(79)N(5)H(1) 0.12 D(3)S(2)L(2) K(3)Q(3) 394 L G(49)I(13)L(15) 0.12 A(16)V(4)C(1) 416 P P(62)A(10)T(4) 0.12 K(6)N(4)S(5) V(4)G(1) 402 C C(71)S(4)G(7) 0.13 F(7)A(8) 417 Y E(42)F(37)Y(9) 0.14 W(2)H(3)L(1) Fig. 7. A possible active surface on the chain 2fbmA. The larger cluster it belongs to is shown in blue. C(1)N(1)A(1) 421 G G(87)A(6)R(3) 0.14 H(1)S(1) The residues belonging to this surface ”patch” are listed in Table 303 S P(79)Q(2)S(9) 0.15 6, while Table 7 suggests possible disruptive replacements for these A(2)N(1)T(3) residues (see Section 3.6). H(1)E(1) 475 V A(52)V(36)T(6) 0.15 Table 6. S(2)L(2) res type substitutions(%) cvg 327 D D(76)N(9)K(1) 0.17 403 D D(97)H(1)T(1) 0.00 R(3)S(6)E(1) 307 N N(97)P(1)H(1) 0.02 A(1) 341 F F(91)Y(8) 0.02 335 S T(63)I(1)S(20) 0.17 494 K K(91)R(6)L(1) 0.02 H(2)R(5)G(2) W(1) A(2)K(1)C(1) 382 P P(93)I(3)V(1) 0.03 343 C A(61)S(24)T(2) 0.17 L(1)M(1) C(9)L(1)V(1) 389 G G(97)N(1)S(1) 0.03 388 N S(4)N(69)E(1) 0.18 460 G G(95)N(1)Q(1) 0.04 H(14)D(3)Q(5) R(1)H(1) A(2) 424 P P(80)A(14)N(2) 0.05 532 I F(73)A(6)I(2) 0.18 L(2)S(1) Y(6).(4)T(3) 346 D D(90)N(3)Y(2) 0.06 Q(1)R(1)L(1) A(2)E(1)S(1) 309 L L(62)V(1)I(15) 0.19 336 A G(82)H(1)A(14) 0.07 F(9)M(8)A(2) S(2) 405 V I(20)V(43)R(30) 0.19 308 A A(83)P(4)S(6) 0.08 A(3)L(1) V(1)F(2)T(2) 299 L L(70)I(18)F(8) 0.20 381 K K(85)C(1)R(3) 0.09 V(2) Q(3)V(2)M(1) 345 L A(42)N(14)G(9) 0.20 H(2)L(1) T(1)L(12)F(2) 386 S A(76)V(10)M(2) 0.09 I(6)D(1)Q(7) G(2)S(5)C(1) V(1)E(1) D(1) 429 S S(40)T(40)A(2) 0.20 425 D G(36)E(16)D(38) 0.10 L(9)H(2)C(1) A(6)P(1)S(1) F(2)I(1) 512 E E(81)A(3)M(5) 0.10 continued in next column continued in next column

5 Table 6. continued Table 7. continued res type substitutions(%) cvg res type disruptive 333 L V(51)L(27)I(16) 0.21 mutations M(2)R(2) 429 S (KR)(Q)(E)(H) 528 S G(56)A(20)K(1) 0.21 333 L (Y)(R)(T)(H) S(12)R(5).(3) 528 S (KR)(FWH)(Y)(EQM) V(1) 392 I (YR)(T)(H)(KE) 392 I L(30)I(21)A(18) 0.22 300 S (FWR)(K)(M)(YH) V(24)M(1)F(3) 398 I (R)(Y)(H)(TK) 300 S N(76)D(4)S(14) 0.23 H(2)R(1)Q(1) Table 7. Disruptive mutations for the surface patch in 2fbmA. 398 I L(68)F(3)I(19) 0.25 V(5)M(2)T(1) 3 NOTES ON USING TRACE RESULTS Table 6. Residues forming surface ”patch” in 2fbmA. 3.1 Coverage Trace results are commonly expressed in terms of coverage: the resi- Table 7. due is important if its “coverage” is small - that is if it belongs to res type disruptive some small top percentage of residues [100% is all of the residues mutations in a chain], according to trace. The ET results are presented in the 403 D (R)(K)(FW)(H) form of a table, usually limited to top 25% percent of residues (or 307 N (Y)(T)(E)(FWH) to some nearby percentage), sorted by the strength of the presumed 341 F (K)(E)(Q)(D) evolutionary pressure. (I.e., the smaller the coverage, the stronger the 494 K (Y)(T)(SCG)(FW) pressure on the residue.) Starting from the top of that list, mutating a 382 P (Y)(R)(H)(T) couple of residues should affect the protein somehow, with the exact 389 G (R)(KE)(FWH)(M) effects to be determined experimentally. 460 G (E)(D)(FW)(KYR) 424 P (YR)(H)(T)(KE) 3.2 Known substitutions 346 D (R)(H)(FW)(K) One of the table columns is “substitutions” - other amino acid types 336 A (KE)(R)(YQ)(D) seen at the same position in the alignment. These amino acid types 308 A (KR)(E)(Y)(Q) may be interchangeable at that position in the protein, so if one wants 381 K (Y)(T)(FW)(S) to affect the protein by a point mutation, they should be avoided. For 386 S (R)(K)(H)(Q) example if the substitutions are “RVK” and the original protein has 425 D (R)(H)(FW)(K) an R at that position, it is advisable to try anything, but RVK. Conver- 512 E (H)(Y)(FW)(R) sely, when looking for substitutions which will not affect the protein, 461 L (R)(Y)(T)(H) one may try replacing, R with K, or (perhaps more surprisingly), with 302 R (T)(Y)(D)(CG) V. The percentage of times the substitution appears in the alignment 394 L (R)(Y)(H)(KE) is given in the immediately following bracket. No percentage is given 416 P (YR)(H)(E)(K) in the cases when it is smaller than 1%. This is meant to be a rough 402 C (K)(ER)(Q)(D) guide - due to rounding errors these percentages often do not add up 417 Y (K)(Q)(E)(R) to 100%. 421 G (E)(K)(R)(D) 303 S (R)(K)(H)(FW) 3.3 Surface 475 V (R)(K)(YE)(H) To detect candidates for novel functional interfaces, first we look for 327 D (FWR)(H)(Y)(CG) residues that are solvent accessible (according to DSSP program) by 2 335 S (KR)(FW)(EQH)(M) at least 10A˚ , which is roughly the area needed for one water mole- 343 C (R)(K)(E)(H) cule to come in the contact with the residue. Furthermore, we require 388 N (Y)(FWH)(T)(R) that these residues form a “cluster” of residues which have neighbor 532 I (R)(Y)(TH)(E) within 5A˚ from any of their heavy atoms. 309 L (YR)(T)(H)(KE) Note, however, that, if our picture of protein evolution is correct, 405 V (Y)(E)(R)(K) the neighboring residues which are not surface accessible might be 299 L (R)(Y)(T)(KEH) equally important in maintaining the interaction specificity - they 345 L (R)(Y)(H)(T) should not be automatically dropped from consideration when choo- continued in next column 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

6 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˚ . 3.5 Annotation COVERAGE

If the residue annotation is available (either from the pdb file or V from other sources), another column, with the header “annotation” 100% 50% 30% 5% appears. Annotations carried over from PDB are the following: site (indicating existence of related site record in PDB ), S-S (disulfide bond forming residue), hb (hydrogen bond forming residue, jb (james bond forming residue), and sb (for salt bridge forming residue).

3.6 Mutation suggestions V

Mutation suggestions are completely heuristic and based on comple- RELATIVE IMPORTANCE 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 Fig. 8. Coloring scheme used to color residues by their relative importance. properties: small [AV GST C], medium [LP NQDEMIK], large [W F Y HR], hydrophobic [LP V AMW F I], polar [GT CY ]; posi- tively [KHR], or negatively [DE] charged, aromatic [W F Y H], by descending size): red, blue, yellow, green, purple, azure, tur- long aliphatic chain [EKRQM], OH-group possession [SDET Y ], quoise, brown, coral, magenta, LightSalmon, SkyBlue, violet, gold, and NH2 group possession [NQRK]. The suggestions are listed bisque, LightSlateBlue, orchid, RosyBrown, MediumAquamarine, according to how different they appear to be from the original amino DarkOliveGreen, CornflowerBlue, grey55, burlywood, LimeGreen, acid, and they are grouped in round brackets if they appear equally tan, DarkOrange, DeepPink, maroon, BlanchedAlmond. disruptive. From left to right, each bracketed group of amino acid The colors used to distinguish the residues by the estimated types resembles more strongly the original (i.e. is, presumably, less evolutionary pressure they experience can be seen in Fig. 8. disruptive) These suggestions are tentative - they might prove disrup- tive to the fold rather than to the interaction. Many researcher will 4.3 Credits choose, however, the straightforward alanine mutations, especially in 4.3.1 Alistat alistat reads a multiple sequence alignment from the the beginning stages of their investigation. file and shows a number of simple statistics about it. These stati- stics include the format, the number of sequences, the total number 4 APPENDIX of residues, the average and range of the sequence lengths, and the 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 • type amino acid type seq” is calculated by finding the maximum pairwise identity (best 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 2 ble if the DSSP program finds it exposed to water by at least 10A˚ , • 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 more than one residue colored according to this hierarchy (ordered http://www.cmbi.kun.nl/gv/dssp/descrip.html.

7 4.3.4 HSSP Whenever available, report maker uses HSSP ali- Evolution-Entropy Hybrid Methods for Ranking of Protein Residues gnment as a starting point for the analysis (sequences shorter than by Importance” J. Mol. Bio. 336: 1265-82. For the original version 75% of the query are taken out, however); R. Schneider, A. de of ET see O. Lichtarge, H.Bourne and F. Cohen (1996). ”An Evolu- Daruvar, and C. Sander. ”The HSSP database of protein structure- tionary Trace Method Defines Binding Surfaces Common to Protein sequence alignments.” Nucleic Acids Res., 25:226–230, 1997. Families” J. Mol. Bio. 257: 342-358. report maker itself is described in Mihalek I., I. Res and O. http://swift.cmbi.kun.nl/swift/hssp/ Lichtarge (2006). ”Evolutionary Trace Report Maker: a new type 4.3.5 LaTex The text for this report was processed using LATEX; of service for comparative analysis of .” Bioinformatics Leslie Lamport, “LaTeX: A Document Preparation System Addison- 22:1656-7. Wesley,” Reading, Mass. (1986). 4.6 About report maker 4.3.6 Muscle When making alignments “from scratch”, report report maker was written in 2006 by Ivana Mihalek. The 1D ran- maker uses Muscle alignment program: Edgar, Robert C. (2004), king visualization program was written by Ivica Res.ˇ report maker ”MUSCLE: multiple sequence alignment with high accuracy and is copyrighted by Lichtarge Lab, Baylor College of Medicine, high throughput.” Nucleic Acids Research 32(5), 1792-97. Houston. http://www.drive5.com/muscle/ 4.7 Attachments 4.3.7 Pymol The figures in this report were produced using The following files should accompany this report: Pymol. The scripts can be found in the attachment. Pymol is an open-source application copyrighted by DeLano Scien- • 2fbmA.complex.pdb - coordinates of 2fbmA with all of its tific LLC (2005). For more information about Pymol see interacting partners http://pymol.sourceforge.net/. (Note for Windows • 2fbmA.etvx - ET viewer input file for 2fbmA users: the attached package needs to be unzipped for Pymol to read • 2fbmA.cluster report.summary - Cluster report summary for the scripts and launch the viewer.) 2fbmA 4.4 Note about ET Viewer • 2fbmA.ranks - Ranks file in sequence order for 2fbmA Dan Morgan from the Lichtarge lab has developed a visualization • 2fbmA.clusters - Cluster descriptions for 2fbmA tool specifically for viewing trace results. If you are interested, please • 2fbmA.msf - the multiple sequence alignment used for the chain visit: 2fbmA http://mammoth.bcm.tmc.edu/traceview/ • 2fbmA.descr - description of sequences used in 2fbmA msf The viewer is self-unpacking and self-installing. Input files to be used • 2fbmA.ranks sorted - full listing of residues and their ranking with ETV (extension .etvx) can be found in the attachment to the for 2fbmA main report. • 2fbmA.2fbmC.if.pml - Pymol script for Figure 5 4.5 Citing this work • 2fbmA.cbcvg - used by other 2fbmA – related pymol scripts The method used to rank residues and make predictions in this report • 2fbmA.2fbmB.if.pml - Pymol script for Figure 6 can be found in Mihalek, I., I. Res,ˇ O. Lichtarge. (2004). ”A Family of

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