An#body (IgG): Analyze the 3D structure of immunoglobulin IgG by assembling this paper model Paper model Based on X-­‐ray structure (PDB ID: 1igt) Contents of ac#vity packet: Paper model template: pages 2-­‐14 Assembling instruc@ons: pages 15 -­‐28 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Light Chain A | Domain 1 la 1F 1G lc lb lb 1B 1A NH3+ la AL1 ld 1E lc 1D le lf le 1H 1C ld AL2 lg 1I lf lh lg Chain A 2 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Light Chain A | Domain 2 li 2E lj lj AL3 ll Chain A lm 2C ll ln lm 2F lk 2D lk li 2B 2A lh AL AL4 COO-­‐ 2G ln 3 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy Chain B | Domain 1 ha he hd 1D 1B hb hd 1E 1G hc hc hb 1F 1A NH3+ ha BH1 hg hf 1H hf 1C he BH2 hh 1I hg hi hh Chain B 4 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy Chain B | Domain 2 hj 2A hk BH3 hl 2E hm hl hn hm ho 2C hn hp ho BH4 2F hk 2D hj 2B hi BH hq 2G hp BHA BHB hr hq Chain B 5 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy Chain B | Domain 3 hs 3A hr BHC ht hu ht 3B hs BH5 hv 3E hw hv 3D hu hx 3C hy hx 3F hw hz 3G hy haa hz Chain B BH6 6 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. 4E hab hac hac 4A hab BH7 had haa 4B Heavy Chain B |Domain 4 4D haf 4C hae 4G hag hag BH8 COO-­‐ haf 4F hae had Chain B 7 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Light Chain C | Domain 1 la 1F 1G lc lb lb 1B 1A NH3+ la CL1 ld 1E lc 1D le lf le 1H 1C ld CL2 lg 1I lf lh lg Chain C 8 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Light Chain C | Domain 2 li 2E lj lj CL3 ll Chain C lm 2C ll ln lm 2F lk 2D lk li 2B 2A lh CL CL4 COO-­‐ 2G ln 9 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy Chain D | Domain 1 ha he hd 1D 1B hb hd 1E 1G hc hc hb 1F 1A NH3+ ha DH1 hg hf 1H hf 1C he DH2 hh 1I hg hi hh Chain D 10 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy Chain D | Domain 2 hj 2A hk DH3 hl 2E hm hl hn hm ho 2C hn hp ho DH4 2F hk 2D hj 2B hi DH hq 2G hp DHA DHB hr hq Chain D 11 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy Chain D | Domain 3 hs 3A hr DHC ht hu ht 3B hs DH5 hv 3E hw hv 3D hu hx 3C hy hx 3F hw hz 3G hy haa hz Chain D DH6 12 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. 4E hab hac hac 4A hab DH7 had haa 4B Heavy Chain D | Domain 4 4D haf 4C hae 4G hag hag DH8 COO-­‐ haf 4F hae had Chain D 13 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. DHB DHC DHA CL2 DH8 DH2 CL4 DH4 DH6 DH AL2 AL4 BH BH2 BHC BH4 BHB BH6 BHA DH7 BH7 DH5 BH5 DH3 BH3 DH1 BH1 CL AL CL3 AL3 CL1 AL1 BH8 Disulfide Linkages 14 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. An#body Model: Overview Light Chain A
Light Chain C
The An@body consists of 4 chains: •  2 light chains: A, C Each light chain is made up of 2 immunoglobulin domains •  2 heavy chains: B, D Each heavy chain is made up of 4 immunoglobulin domains Heavy Chain B
Heavy Chain D
15 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. An#body Model: Overview Assembly Overview: •  Each page from 2 to 13 contains elements to build 1 domain •  You will fist create the domains and then assemble the chains •  The an@body structure is stabilized by 17 disulfide linkages colored yellow on the image to the right •  The stripes in the paper model template, on page 14, represent these linkages •  Crea@ng the disulfide linkages is the last step in making each Ig domain and finally the complete an@body molecule. 16 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Prepara#on For each page from 2 to 13 : •  Cut out each strip along the do\ed lines •  Note that the shaded regions labeled ho, hk, etc. should be included in the strip •  When crea@ng the domains, work on one page at a @me to avoid mixing up the strips. Make sure these shaded regions remain a\ached to the strips 17 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Immunoglobulin Domains For each page from 2 to 13 : Step 1 Create the representa@on of the polymer backbone •  Tape together the strips into a long strand •  The labels la, lb, etc. on the grey background should be hidden underneath the labels on the colored background Find matching labels and tape together hiding the hatched grey label. Polymer backbones for all 12 domains. 18 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Immunoglobulin Domains Step 2 Create Immunoglobulin Domains The solid black arrows labeled 1A, 1B, 1C, 1D etc. represent the beta strands. Create the beta strand clusters by taping together the strands side by side. For each domain, make sure that: 1.  The le[ers on the rectangular labels are in alphabe#cal order, for example 4A-­‐4B-­‐4C-­‐4D, or 4E-­‐4F-­‐4G. Note: Don’t connect the strips if there isn’t any grey area in between them. 2.  The grey area between the strands is aligned. (See next page) 3.  The beta strand arrows point up and down alternately (See next page) Note: No grey area. Stop here and begin a new cluster star@ng with 4E through 4G. The le\ers inside the rectangular labels follow alphabe@cal order Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. 19 Immunoglobulin Domains The beta strand arrows point up and down alternately The gray area between strands is aligned. All 12 domains assembled 20 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Heavy and Light Chains Assembling the 4 polymer chains: Light Chains A and C: •  Tape together the 2 domains of each chain at the label lh; Heavy Chains B and C: •  Tape together the 4 domains of each chain at the labels hi, hr and haa Align and tape; Grey labels should be hidden underneath the colored ones. Note: Follow the chain labels to make sure you are connec@ng the domains from the proper chains. 21 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Light and Heavy Chains 2 light and 2 heavy polymer chains assembled 22 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Disulfide Linkages The Structure of An@body is stabilized by 2 types of disulfide linkages: •  12 Intradomain linkages stabilize each of the 12 immunglobulin domains. •  5 Interchain linkages hold the different chains together •
Intradomain Linkages (1 for each domain) Interchain Linkages The 17 strips on page 14 represent the disulfide linkages •  Cut out all strips on the do\ed lines •  Fold down each strip on both hatched regions. 23 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Disulfide Linkages Create the Intradomain Disulfide Linkages: For each domain: •  Find corresponding labels on the domains (round, marked AL1 and AL2) and on the linkages •  Align the strip label underneath the domain’s round label and tape Corresponding labels The linkage strip labels are aligned and taped underneath the round labels 24 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Disulfide Linkages Note: Before you start the final step, it might be a good idea to align the light and heavy chains. Use the images on pages 15 (overview sec@on) for help with domain and chain placement in the model. This step will help with label iden@fica@on and proper placement of the interchain linkages. 25 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Disulfide Linkages Create the interchain disulfide linkages: •  Chains A and B: Iden@fy the round, do[ed labels AL and BH on the model and connect them with corresponding disulfide strip. The printed sides of the labels should face each other. •  Repeat for chains C and D (labels CL and DH) Chains are connected at round do\ed AL and BH labels. The printed sides of the labels face each other. 26 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Disulfide Linkages Create the interchain disulfide linkages: Chains B and D: •  Iden@fy the do\ed round labels BHA–DHA BHB–DHB BHC–DHC on the model and on the disulfide strips •  Tape the printed side of the hatched regions on top of the do\ed round labels. Chains B and D are connected at round do\ed labels. The printed sides of the labels face each other. 27 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014. Finished Model Sugges#on: This model can be made as a group project. Assign each domain to a different individual. The 12 different domains in the an@body can be assembled together and all the disulfide bridges put in place as a group. 28 Developed as part of the RCSB Collabora@ve Curriculum Development Program 2014.