1. Electrostatic Potential on HLA-DPB1*1701 and HLA-DPB1*0401: Implications for Putative Mechanism of Chronic Beryllium Disease Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, WV James A. Snyder, Ainsley Weston, Sally S. Tinkle and Eugene Demchuk

2. Objective to understand the physical-chemical reasons for beryllium toxicity in Chronic Beryllium Disease (CBD)

3. Periodic Table What makes Be toxic?

4. The Test Hypothesis Be 2+ binds to HLA-DP and changes the specificity of molecular recognition Goal: To identify and test the Be 2+ binding mode

5. Experimental Evidence Amicosante, M., Sanarico, N. Berretta, F., Arroyo, J., Lombardi, G., Lechler, R., Colizzi, V. and Saltini, C. Beryllium Binding to HLA-DP Molecule Carrying the Marker of Susceptibility to Berylliosis Glutamate  69. Human Immunology, 62, 686-693 (2001) Fontenot, A.P., Torres, M., Marshall, W.H., Newman, L.S. and Kotzin, B.L. Beryllium Presentation to CD4 + T Cells Underlies Disease-Susceptibility HLA-DP Alleles in Chronic Beryllium Disease. Proc. Natl. Acad. Sci. USA, 97, 12717-12722 (2000)

6. MHC class II cell membrane  

7. Crystal Structure of Human Class II MHC Proteins in the Protein Data Bank (PDB) HLA-DR1 1AQDHLA-DR1 (DRA, DRB1 0101) Human MHC Class II complexed with endogenous peptide 1DLHHLA-DR1 (DRA, DRB1 0101) Human Class II histocompatibility protein (extracellular domain) 1FYTHLA-DR1, DRB1 0101 1HQRDRA*0101 + DRB5*0101 1KG0the Epstein-Barr virus Gp42 Protein bound to the MHC Class II HLA-DR1 1SEBcomplex of the Human MHC Class II Glycoprotein HLA-DR1 and the bacterial superantigen SEB HLA-DR2 1BX2HLA-DR2 (DRA*0101, DRB1*1501) complexed with a peptide from Human myelin basic protein 1FV1immunodominant peptide from myelin basic protein in different registers by two HLA-DR2 alleles HLA-DR3 1A6Aclip bound to HLA-DR3 HLA-DR4 1D5MHLA-DR4 complexed with peptide and SEB 1D5XHLA-DR4 complexed with dipeptide mimetic and SEB 1D5ZHLA-DR4 complexed with peptidomimetic and SEB 1D6EHLA-DR4 complex with peptidomimetic and SEB 1J8HHLA-DR1, DRA + HLA-DR4, DRB1*0401 2SEBHLA-DR4 complexed with a peptide from Human collagen II HLA-DQ8 1JK8Human insulin peptide-HLA-DQ8 complex HLA-DM 1HDMhistocompatibility antigen HLA-DM

8. of HLA-DPB1*1701 with  chain of HLA-DR2 (PDB: 1FV1B) 10 20 30 40 *1701 RATPENYVHQLRQECYAFNGTQR - - FLERYIYNREEFVRFDS 1FV1  GDTRPRFLQQDKYECHFFNGTERVRFLHRDIYNQEEDLRFDS 10 20 30 40 50 60 70 80 *1701 DVGE FRAVTELGRPDEDYWNSQKDI LEEERAVPDRMCRHN 1FV1  DVGEYRAVTELGRPDAEYWNSQKDFLEDRRAAVDTYCRHN 50 60 70 80 90 100 110 120 *1701 YELDEAVTLQRRVQPKVNVSPSKKGPLQHHNLLVCHVTDF 1FV1  YGVGESFTVQRRVEPKVTVYPARTQTLQHHNLLVCSVNGF 90 100 110 120 130 140 150 160 *1701 YPGSIQVRWFLNGQEET AGVVSTNLIRNGDWTFQ ILVMLE 1FV1  YPGSIEVRWFRNS QEEKAGVVSTGLIQNGDWTFQTLVMLE 130 140 150 160 170 180 190 200 *1701 MTPQQGDVY ICQVEHTSLDSPVTVEWKAQSDSAQSKTLTG 1FV1  TVP RS GEVYTCQVEHPSVTSPLTVEWRA 170 180 190 Sequence Alignment

9. Molecular Dynamics…

10. of  -carbon atom coordinates relative to starting (crystallographic) positions at 1 Pico-second intervals Total Deviation

11. Total Charge on HLA-DP proteins DR Template Protein: -14  HLA-DPA1*01031/B1*0401: -17/-3 HLA-DPA1*01031/B1*1701: -17/-9 Be 2+

12. CBD Susceptibility Odds and the Charge on  -chain of HLA-DP HLA-DPB1 Odds Charge *1701-9 *0601-9 *0901, *1001 -9, -9 *1901-8 *0201-7 *0401-3

13. 1 40 *1701 RATPENYVHQ LRQECYAFNG TQRFLERYIY NREEFVRFDS *0401 RATPENY LFQGRQECYAFNG TQRFLERYIY NREEFARFDS 41 -1-1 -2 80 *1701 DVGEFRAVTE LGRPDEDYWN SQKDILEEERAVPDRMCRHN *0401 DVGEFRAVTE LGRPAAEYWN SQKDILEEKRAVPDRMCRHN 81 -1-1 120 *1701 YELDEA VTLQRRVQPKVNVS PSKKGPLQHH NLLVCHVTDF *0401 YELGGPMTLQRRVQPRVNVS PSKKGPLQHH NLLVCHVTDF 121 160 *1701 YPGSIQVRWF LNGQEETAGV VSTNLIRNGD WTFQILVMLE *0401 YPGSIQVRWF LNGQEETAGV VSTNLIRNGD WTFQILVMLE 161 188 *1701 MTPQQGDVY I CQVEHTSLDS PVTVEWKA *0401 MTPQQGDVYT CQVEHTSLDS PVTVEWKA Sequence Alignment of HLA-DPB1*1701 with HLA-DPB1*0401

14.  -chain Class II MHC antigen-binding Pocket  -chain

15. Poisson-Boltzmann Equation 

16. Finite Difference PB Methods  using divergence theorem:

17. HLA-DPA1*01031/B1*0401 (model) Electrostatic Potential Surface Map

18. HLA-DPA1*01031/B1*1701 (model) Electrostatic Potential Surface Map

19. *1701 MEP Movie

20. *0401 MEP Movie

21. 1701 MEP Movie

22. Conclusion Our results are consistent with the hypothesis that glutamic acid at position 69 of the  -chain of HLA- DP may be a determinant of susceptibility to Chronic Beryllium Disease

23. James A. Snyder Eugene Demchuk Ainsley Westongenetics Sally S. Tinkleimmunology People Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, WV computational studies

24. Future Goal: To model interactions of Be 2+ with HLA- DP proteins