Guide t Using the Rubric t Scre the Caspase-3 Pre-Build Mdel fr Science Olympiad 2011-2012 Natinal Cmpetitin These instructins are t help the event supervisr and scring judges use the rubric develped by the MSOE Center fr BiMlecular Mdeling when scring the 2011-2012 Science Olympiad Natinal Pre-Build mdels f Caspase-3, based n 1i3.pdb. Each categry n the rubric is addressed within these instructins and is accmpanied by a shrt descriptin and picture, if apprpriate. Overview f the Mlecule: A caspase-3 mnmer is cmprised f tw chains. Chain A (clred light blue in the figure t the left) is lnger than Chain B (clred purple). Impunded tber mdels shuld have tw clrs t differentiate the chains. A different clring scheme fr caspase-3 is shwn in the figure t the right. In this clr scheme, helices are highlighted in magenta and sheets are clred yellw. The N and C termini f the tw chains are clred blue/light blue and red/marn, respectively.
1. Blue Caps n N-terminal Amin Acids (Ser148 n Chain A; Ser310 n Chain B) (0.5 pts; 0.25 pts fr each N-terminus f the tw chains) Please nte: Teams using nn-tber mdels may have ther methds f indicating the N- and C-termini aside frm the blue and red plastic end caps prvided when a team purchased a prebuild kit frm 3D Mlecular Designs. Chice f material is nt part f the scring; hwever, the team s 3x5 nte card shuld include an explanatin f selected materials fr the demarcatin f the N- and C-termini t eliminate guesswrk n the part f the event supervisr. T receive credit fr this feature, the mdel shuld have tw blue caps, indicating the N-terminus f bth Chain A and Chain B. Please see the figures and explanatin belw fr the crrect psitining f these blue end caps. In the figure n the left, the A chain is clred cyan and the B chain is clred purple. The figure n the right has the prtein in the same rientatin, with a different clring scheme. The helices are clred magenta and the sheets are clred yellw. N-Terminus f Chain A N-Terminus f Chain B Chain A (the lnger f the tw chains; clred cyan in the figure n the left): The blue cap shuld be psitined befre β-strand #1, fllwed by anther β-strand that frms the 2 nd strand in the central β-sheet. The central β-sheet is cmpsed f strands frm bth Chain A and Chain B. Chain A cntributes strands 1-4 f the 6 stranded sheet. The final strand (strand #4) in Chain A s prtin f the central beta sheet shuld lead int the C-terminus f Chain A. T differentiate between the N and C-terminus regins f the prtein, fllw the tber backbne frm the N-terminus. If the path frm the N-terminus includes a shrt β-strand, fllwed by a lnger β-strand that frms a middle strand f the central β-sheet, then the cap is crrectly psitined. (0.25 pts) If the backbne leads frm the end cap t a shrt β-strand and then t a strand that frms the uter edge f Chain A s cntributin t the central β-sheet (the middle strand f the entire β-sheet), then the mdel shuld nt receive pints fr this feature, fr this is the C-terminal end f the prtein and shuld have the red end cap psitined here. Chain B (the shrter f the tw chains; clred purple in the figure t the abve left): The blue end cap shuld be psitined n the end that has bth a shrt helix and a shrt beta strand. (0.25 pts) If the blue end cap is psitined near a beta strand nly, then the mdel des nt receive pints fr this feature.
2. Red Caps n C-terminal Amin Acids (Thr296 n Chain A; Tyr401 n Chain B) (0.5 pts; 0.25 pts fr each C-terminus f the tw chains) T receive credit fr this feature, the mdel shuld have tw red caps, indicating the C-terminus f bth Chain A and Chain B. Please see the figures and explanatin belw fr the crrect psitining f these red end caps. In the figure n the left, the A chain is clred cyan and the B chain is clred purple. The figure n the right has the prtein in the same rientatin, with a different clring scheme. The helices are clred magenta and the sheets are clred yellw. C-Terminus f Chain A C-Terminus f Chain B Chain A (the lnger f the tw chains; clred cyan in the figure n the left): The red cap shuld be psitined at the end the shrtest β-strand f Chain A. This strand shuld extend ut frm the middle f the central β-sheet. The central β-sheet is cmpsed f strands frm bth Chain A and Chain B. Chain A cntributes strands 1-4 f the 6 stranded sheet. The final strand (strand #4) in Chain A s prtin f the central beta sheet shuld lead int the C-terminus f Chain A. T differentiate between the N- and C-terminus regins f the prtein, fllw the tber backbne back frm the C-terminus. If the path frm the C-terminus includes a shrt strand, fllwed by a lnger β-strand that frms the uter edge f the central β-sheet, then the cap is crrectly psitined. (0.25 pts) If the backbne leads frm the end cap t a strand that frms the a middle strand f the central β-sheet, then the mdel shuld nt receive pints fr this feature, fr this is the beginning f the prtein and shuld have the blue end cap psitined here. Chain B (the shrter f the tw chains; clred purple in the abve figure t the left): The red end cap shuld be psitined n the end that has beta strands. If the red end cap is psitined near a shrt beta strand and a shrt helix, then the mdel des nt receive pints fr this feature. (0.25 pts)
3. Mdel has 6 alpha helices accrding t Jml selectin criteria (0.5 pts per helix fr a ttal f 3 pints) T receive these pints, there shuld be 6 helices within the mdel. Please see figure t the right fr the crrect lcatin f these helices. (Helices are clred magenta n the mdel and in the figure t the right.) Deduct 0.5 pints fr each extra helix. Fr example, if a mdel has 7 helices, then the mdel wuld receive 2.5 pts rather than the full 3 pints. 4. Alpha helices are right-handed (0.5 pts each; ttal f 3 pts) Alpha helices are right-handed. Check each alpha helix in the mdel t cnfirm that the helix is right-handed. Fr each right-handed helix, the mdel shuld receive 0.5 pints, fr a ttal f 3 pints if all six helices are crrect. If the mdel nly has 5 helices, and all five are right-handed, then the mdel wuld nly receive 2.5 pints. T determine if the helix is right-handed, find ne f the ends f the helix and imagine that the helix is a spiral staircase. Pretend that yu are climbing that staircase and yu need t have a hand-rail and the helix is the hand-rail, which is always n the utside edge f the staircase. If yu wuld put yur right hand n the tber as yu g up the staircase, yu have a right-handed helix. If yu wuld put yur left hand n the tber, yu have a left-handed helix and the mdeled helix wuld nt receive credit. Left-handed vs righthanded helices 5. Alpha helices are the crrect length. (0.5 pts each; ttal f 3 pts) Each helix that is f the crrect length shuld receive 0.5 pt, fr a ttal f 3 pts. Please nte that the helices are numbered sequentially, with the first helix identified as the ne clsest t the N- terminus f that chain. Chain A Helix #1 is 15 amin acids lng. The helix shuld have 4-5 turns. Helix #2 is 13 amin acids lng. The helix shuld have 4-5 turns. Helix #3 is 8 amin acids lng. This helix shuld have 2-3 turns. Chain B Helix #1 is 4 amin acids lng. The helix shuld have 1-2 turns. Helix #2 is 15 amin acids lng. The helix shuld have 4-5 turns. Helix #3 is 17 amin acids lng. The helix shuld have 5-6 turns.
6. Mdel has 11 β-strands (0.5 pts per β-strand fr a ttal f 5.5 pints) PLEASE NOTE: Fr features #6 (number f beta-strands) and #7 (number f sheets), please use the images prvided within this rubric t scre the mdels, rather than the plaster mdel that accmpanied the materials prvided fr the natinal turnament. Due t a technical errr, the mdel has the incrrect number f betastrands highlighted. This errr des nt impact any ther part f the scring rubric. The three-dimensinal shape f the prtein is accurately reflected in the mdel. Therefre, the plaster mdel can be used t scre the student-cnstructed mdels fr all ther features. T receive these pints, there shuld be 11 β-strands within the mdel. Chain A (the lnger f the tw chains) has 8 strands. (0.5 pts each fr a ttal f 4 pts ttal) Fur f the strands are in the center beta sheet and these shuld be the lngest f the strands within Chain A. Three f the strands frm a smaller beta sheet (highlighted in the image belw). These strands shuld be shrter than the strands that create the central beta sheet. There is ne small beta strand near the C-terminus. This strand is nly a cuple amin acids in length and is the smallest strand n Chain A. Chain B (the shrter f the tw chains) has 3 strands. (0.5 pts each fr a ttal f 1.5 pts) Tw f the strands are in the central beta sheet, aligned with the strands frm Chain A. These strands shuld be the lngest strands within Chain B. There is ne small beta strand lcated after the helix near the N-terminus. This strand is nly a cuple amin acids in length and is the smallest strand n Chain B. Please see figures belw fr the crrect lcatin f these beta strands. (β-strands are clred yellw.) 6 strands are lcated in the middle f the prtein. There is a shrt strand lcated near the C- terminus f the A chain (highlighted red in the image belw) and the N-terminus (highlighted cyan in the image belw) f the B Chain. Three strands are lcated n the edge f the prtein.
β-strands need t be clearly distinguishable frm lps; there may be sme slight zig-zag flding f the tber t indicate the up-and-dwn psitining f the amin acids. Alternately, teams might clr-cde their beta strands t distinguish them frm lps r write n the tber indicating the lcatin f the β-strands. The event supervisr shuld nt have t guess what a beta-strand is within the mdel. If there are mre than 11 β-strands in this mdel, 0.5 pts shuld be deducted fr each extra strand. Fr example, if the mdel has 12 β-strands, the mdel shuld receive 5 pints, rather than the full 5.5 pints. 7. Mdel has 2 β-sheets (2 pts fr each sheet fr a ttal f 4 pints) This prtein has tw β-sheets arranged frm the 11 β-strands. A β-sheet is cmpsed f three r mre β-strands lying parallel t each ther, thugh the plane f the β-sheet may be twisted instead f flat. β-sheet #1 (the large ne in the center) is frmed by strands 1, 2, 3 and 7 frm Chain A and strands 2 and 3 frm Chain B. (Strands are numbered in the rder f appearance fr each chain, based n the prximity t the respective N-terminus.) There are a ttal f 6 strands frming this center β-sheet. In the images t the right, Chain A is clred cyan and Chain B is clred purple. The sheets are clred yellw. (2 pts) β-sheet #2 is frmed by strands 4,5 and 6 f Chain A, which is psitined near helices #2 and #3 n Chain A. (2 pts) If the mdel has mre than tw β-sheets, then deduct 1 pt fr each additinal sheet. Fr example, if the mdel has three β- sheets, the mdel shuld receive 3 pts, rather than the full 4 pts. Beta Sheet #1 (lcated in the center f the prtein) is cmpsed f 6 strands. Beta Sheet #2 is cmpsed f 3 strands (all frm Chain A).
Items #8-#10 refer t the three-dimensinal shape f the prtein. Layer 1 Layer 2 8. Mdel has three layers (0.75 pts) Layer 3 Mdel can be viewed as having three distinct layers. If yu hld the mdel s that the center beta sheet is lying perpendicular t the flr, there shuld be a layer with tw helices, a center beta sheet and a layer with three helices. The sheet layer (Layer 2 n the image t the right) is sandwiched between the helices layers (Layers 1 and 2 n the image t the right). Please refer t the physical mdel and the figure t the right. If the mdel has all three layers, award 0.75 pts. It the mdel has tw layers, award 0.5 pts. If the mdel has a single layer, award 0.25 pts. If the mdel des nt have any discernable layers, n pints shuld be awarded. 9. The layer with three helices is cmprised f ne helix frm Chain A and tw helices frm Chain B (0.5 pts) The caspase-3 mnmer is cmprised f tw chains (Chains A and B; clred cyan (Chain A) and purple (Chain B) in figure t right). Kits purchased thrugh 3D Mlecular Designs included tw different clred tbers t cnstruct their mdels t illustrate the tw chain feature f this mdel. If a mdel des nt have a clr differentiatin between the chains, please take the time t determine the chain arrangement n the mdel. Chain A (cyan) cntributes a single helix t this helix layer (helix in dark blue). Chain B (purple) cntributes tw helices t this helix layer (helices clred pink). Hld the mdel s that the three helices are n the tp and s that Chain B (clred purple in figure t the right) is lcated n the right-hand side f the mdel (see figure t right fr crrect psitining). T receive credit fr this feature, there shuld be three helices n this layer, tw f which are frm chain B (clred pink n figure t the right) and ne frm Chain A (clred blue in figure t the right). (0.25 pts) Helices shuld be arranged s that the tw helices n chain B (pink in abve figure) are lying next t each ther and the helix frm Chain A lies t ne side. (0.25 pts) If the mdel has the helix frm chain A psitined between helices frm Chain B, the mdel shuld nt receive credit fr this feature.
10. The layer with tw helices is cmprised f tw helices frm Chain A (0.5 pts) The caspase-3 mnmer is cmprised f tw chains (Chains A and B; clred cyan (Chain A) and purple (Chain B) in figure t right). Kits purchased thrugh 3D Mlecular Designs included tw different clred tbers t cnstruct their mdels t illustrate the tw chain feature f this mdel. If a mdel des nt have a clr differentiatin between the chains, please take the time t determine the chain arrangement n the mdel. Hld the mdel s that the tw helices are n the tp and s that Chain B (clred purple in figure t the right) is lcated n the left-hand side f the mdel (see figure t right fr crrect psitining). T receive credit fr this feature, there shuld be tw helices n this layer, bth frm chain A (clred blue n figure t the right). (0.25 pts) Helices shuld be arranged s that the tw helices n chain A (blue in abve figure) are lying next t each ther. (0.25 pts) Chain A (cyan) cntributes bth helices in this layer (helices clred dark blue). If the mdel has mre than tw helices n this half, r that there is a helix frm chain B included in the layer, the mdel des nt receive credit fr this feature. N-terminus B chain (cyan) 11. Three f the end caps are lcated n the same side f the mdel (N and C f B chain, N f A chain) (0.75 pts) Hld the mdel t view the central beta sheet frm the side. The layer f the mdel with the tw helices shuld be n the tp and the layer with the three helices shuld be n the bttm. The N-terminus end caps fr Chains A and B shuld be lcated n the right side. Please see the figure t the right fr crrect rientatin. Three f the end caps shuld be lcated n the right side f the mdel in this rientatin. (0.25 pts per crrectly psitined terminus; fr a ttal f 0.75 pts) The N-terminus f the A chain (blue n figure t the right). The N-terminus f the B chain (cyan n figure t the right). The C-terminus f the B chain (dark red n figure t the right). C-terminus B chain (dark red) N-terminus A chain (blue)
12. The C-terminus f the A chain is lcated n ppsite half f the mdel frm the ther 3 end caps (0.25 pts) Hld the mdel t view the central beta sheet frm the side. The layer f the mdel with the tw helices shuld be n the tp and the layer with the three helices shuld be n the bttm. The N-terminus end caps fr Chains A and B shuld be lcated n the right side. Please see the figure t the right fr crrect rientatin. The C-terminus f the A chain shuld be n the left side f the mdel (red n the figure t the right) (0.25 pts) C-terminus A chain (red) 13. Students submitted a 3x5 card with explanatin f the mdel (0.75 pts) The 3x5 card submitted with the mdel shuld describe the mdel in terms f what additinal features have been added t the mdel s that the judge is nt left guessing what the mdel represents. 14. Functinally Significant Additins (16 pts) Please nte that there are many pssible functinally significant additins that culd be added t the mdel. Fr the Natinal Turnament, each additin is wrth 4 pts; teams may have mre than 4 creative additins, but may nt earn mre than 16 pts in this sectin. If yu are using the electrnic scre sheet, it will autmatically calculate the sum f pints and recrd a maximum f 16 pints. Active site amin acids (Cys285 and His237) (4 pts) Caspase is a prtease, an enzyme that cleaves prtein targets. Enzymes have an active site, which cnsists f specific amin acids that perfrm the catalytic functin f the enzyme. In caspase, the active site cnsists t tw amin acids: His237 and Cys285. T receive credit fr this feature, the mdel must have His237 and Cys285 psitined apprpriately within the prtein. (These sidechains are depicted in green n the image n the next page). (1 pt per amin acid) His237 and Cys285 are bth lcated n chain A (the lnger f the tw chains). His237 shuld be psitined n beta strand #5, the first strand that frms the smaller β-sheet. Cys285 shuld be psitined n the tber near the central beta sheet, where the backbne extends t lead int the C-terminus.
The sidechains must als be psitined apprpriately relative t ne anther If yu hld the mdel with the three helix half n tp and bth N-termini tward the back f the mdel (yu want t lk at the center beta-sheet with the amin acids facing yu), bth His237 and Cys285 shuld be psitined pinting tward the three helix face f the mdel. See figure abve fr crrect rientatin. (1 pt per amin acid) The sidechains shuld nt be pinting twards ne anther r away frm ne anther, but rather upward tward the three helices. N pints shuld be awarded if the sidechains are nt pinting tward the helices. Additin f 2 nd Caspase Mnmer t create functinal dimer (4 pts) Caspase-3 functins as a dimer. Teams may elect t add t their mdel anther caspase mnmer. In this PDB file, Chains C and D cmprise the 2 nd caspase mnmer, as shwn in the figure t the right clred green. Students may elect t represent the additinal capsase mnmer in a variety f ways. Pints shuld be awarded if psitined apprpriately n their mdel f Chains A and B. (2 pts) The 2 nd mnmer f caspase shuld be psitined near the N and C-termini f Chains A and B. Amin acids in the active site are displayed n the additinal mnmer. (2 pts) The active site is cmprised f His237 and Cys285 (see abve ntes abut psitining n amin acids. Chains C and D are identical t Chains A and B, respectively.
Caspase-3 targets (4 pts) Caspase-3 cleaves specific prteins t induce the apptsis f the cell. Teams may include prtein targets f caspase-3 t indicate the enzymatic activity f caspase-3. Target prteins include PARP (ply APD Ribse Plymerase), DFF (DNA fragmentatin factr), nuclear lamins, ICAD (inhibitr f caspase-activated DNase) and PAK2 (P21-activated Kinase 2). Pints shuld be awarded if target prtein is apprpriate and explained by the 3x5 nte card. XIAP bund t Caspase-3 (4 pts) BIR Dmain XIAP is an inhibitr f caspase-3. This prtein (shwn in green n figure t the right) blcks the active site, preventing caspase-3 frm cleaving its target substrates. XIAP is als the prtein where the mutatin in Nic Vlker was identified thrugh genmic sequencing. XIAP has hk, line and sinker regins that lay acrss the prtein, ccupying the active site. (2 pts) The BIR2 dmain (see figure t the right) f the XIAP prtein des nt interact with the active site f caspase- 3. The active site f Caspase-3 is blcked by the hk, line and sinker regins f the XIAP prtein. (2pts) Hk, line and sinker regin f XIAP Pints shuld be awarded if the bund XIAP is present, ccupying the active site and explained n the 3x5 nte card. Amin acids n Caspase-3 that interact with the BIR2 dmain f XIAP (Leu290, Tyr338, Trp340, Phe381^h) (4 pts) The BIR2 dmain f XIAP interacts with caspase-3. Leu290, Tyr338, Trp340, Phe381^h play a rle in binding XIAP Pints shuld be awarded if these amin acids are represented n the mdel in the apprpriate lcatin and rientatin (clred green n the image t the right) Leu290 is lcated n chain A n the backbne that extends away frm the central β-sheet and leads t the C-terminus, just befre the final, shrt β-strand. Tyr338 is lcated n Chain B n the lp between β-strand #2 and helix #2.
Trp340 is lcated n Chain B n the lp between β-strand #2 and helix #2. Phe381^h is lcated n Chain B n the lp between helix #3 and β-strand #3. The armatic rings frm 3 sides f a bx, and Leu pints inward t the bx. Please see figure t the right. The three armatic sidechains (Tyr, Trp and Phe) are clred green. Leu is clred cyan. If all fur amin acids are psitined crrectly, award 4 pts. If three f the fur amin acids are psitined crrectly, award 3 pts. If tw f the fur amin acids are psitined crrectly, award 2 pts. If ne f the fur amin acids is psitined crrectly, award 1 pt. Additinal features that are nt listed abve may be added t the prtein. It is up t the event supervisr t determine whether the additin t the mdel is apprpriate and accurate. In rder t receive pints, the additins must be n the mdel and a cmplete descriptin and explanatin f significance must be included n the 3x5 nte card. A mdel cannt receive mre than 12 pts ttal fr this sectin. 15. Additins t mdel are apprpriate t functin f the prtein (1 pt) T receive credit fr this feature, the creative additins need t be relevant t telling the functinal stry f the prtein. Credit shuld be awarded t thse mdels that meet the fllwing criteria: Mdel has creative additins Mdels that are just the tber will nt receive credit Additins are apprpriate t the functin f the prtein Mdels that have ALL sidechains displayed shuld nt receive credit. If all sidechains are displayed, the team did nt recgnize the significance f a select few amin acids t the enzyme functin. The fcus f adding the amin acids shuld be n the nes that play a rle in the functin f the prtein. Additinally, credit shuld nt be awarded if amin acids utside f thse that play a specific functinal rle in the prtein are displayed.