Project: identify potential ligand binding to the heme-binding site of pHus using DOCK Investigators; Maura J. O Neill Xiao Zhu Kellie Hom hom@nmr.umaryland.edu Angela Wilks awilks@rx.umaryland.edu Alex Mackerell # Tue Apr 1 12:48:14 EST 2008 Created /home/hom/@wilks/@phus_drugscreening received the following three items from Maura O'Neill : -rw-rw-r-- 1 hom hom 128232 Apr 1 13:00 final_181_merged.pdf -rw-rw-r-- 1 hom hom 135168 Apr 1 12:33 final_181_merged.xls a xerox copy of the original purchase order from Maybridge, 168 compounds. #Wed Apr 23 15:01:10 EDT 2008 Created individual gif images for each of 181 compounds: pdftopbm final_181_merged.pdf final_181_merged produced final_181_merged-000001.pbm to final_181_merged-000037.pbm, 38 files in all; used gimp on each to cut out image and saved as gif files. #May 19 finished html: ~/@wilks/@phus_drugscreening/@web_pHus_ligand/pHus_screening.html # Tue Jun 3 13:29:52 EDT 2008 The compounds were from Maybridge and delivered to us in 96-well plates. Maura O'Neill and Kim Burkhard added 1mL of CHCl3 to each well and aliquoted 100 microliter of each in 0.5 mL eppendorfs, dried down the chloroform and gave them to me. I also received on 05/30/08 a file from Maura, describing the correspondance between rack poisitons and identities of compounds, On the packing slip from maybridge, I have put down on each line, the local number names, 001 ... 181, that I made for each compound. I add 300 microliter of D2O to each of R1A2, R1A3, R1A4 and R1A5. Let it sit at room temperature for 1 hour. Take 135 uL of each and combine in a 1.5 mL eppendorf. Add 60uL of 500 mM phosphate in D2O, rack maybridge local STD ==== ========= ===== === R1A2 AW00295SC 001 - R1A3 BTB06287SC 010 - R1A4 BTB15078SC 019 - R1A5 CD10589SC 027 - Received from Maura, 5 mL of pHus in tris buffer. I dialyzed it against pH7 phosphate buffer, sodium 2X (2 liter and 1 liter). Concentration before dialysis: 13 mg/mL, about 0.2 mM Assume that the concentration is still 0.2 mM in the phosphate. Divided it into 5-1mL aliquots 135 uL each of R1A2, R1A3, R1A4 and R1A5, 60 uL of 500 mM phosphate in D2O, 10 uL of pHus, 2 uL of TSP. #Wed Jun 4 10:32:21 EDT 2008 No binding pHus to R1A2(001), R1A3(010), R1A4(019), R1A5(027) I will try 5 uL of pHus for next batch. rack maybridge local STD ==== ========= ===== === R1B1 - R1B2 AW00414SC 002 - R1B3 BTB06700SC 011 - R1C1 - # Jun 9, 2008 rack maybridge local STD ==== ========= ===== === R1C3 BTB07932SC 012 - R1C4 CD04055SC 021 - R1C5 CD11359SC 029 - R1D2 AW00850SC 004 - # Fri Jun 20 09:55:39 EDT 2008 06/19/2008 rack maybridge local STD ==== ========= ===== === R1A6 DSHS00893SC 035 - R1B6 GK00799SC 036 - R1C6 GK00899SC 037 - R1D3 BTB09925SC 013 - 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 06/23/2008 rack maybridge local STD ==== ========= ===== === R1D4 CD05035SC 022 - R1D5 CD11549SC 030 - R1D6 GK01763SC 038 - R1E2 BTB00637SC 005 - 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 06/24/2008 rack maybridge local STD ==== ========= ===== === R1E3 BTB10802SC 014 + R1E4 CD08348SC 023 - R1E5 DFP00365SC 031 - R1E6 GK01800SC 039 + 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 06/25/2008 rack maybridge local STD ==== ========= ===== === R1F2 BTB02067SC 006 - R1F3 BTB11506SC 015 - R1F4 CD08623SC 024 - R1F6 GK02931SC 040 - 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 06/26/2008 rack maybridge local STD ==== ========= ===== === R1E9 HTS07162SC 063 ~ R1F9 HTS07357SC 064 ~ R1G9 HTS07756SC 065 ~ R1H9 HTS07764SC 066 ~ 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 07/01/2008 rack maybridge local STD ==== ========= ===== === R1A10 HTS09664SC 067 - R1B10 HTS10167SC 068 - R1C10 HTS10187SC 069 - R1D10 HTS10813SC 070 ? - 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 07/16/2008 rack maybridge local STD ==== ========= ===== === R1E10 HTS11138SC 071 ~ R1F10 HTS11321SC 072 ~ R1G10 HTS13296SC 073 ~ R1H10 HTS13501SC 074 ~ 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 07/23/2008 rack maybridge local STD ==== ========= ===== === R1A11 JFD00715SC 075 + R1B11 JFD00741SC 076 + R1C11 JFD01500SC 077 - R1D11 JFD01751SC 078 - 300uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). # 07/24/2008 rack maybridge local STD ==== ========= ===== === R1E11 SP01419 167 - R1F11 SP01482 168 - R1G11 SPB01208 169 + ? R1H11 SPB02111 170 - ? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 07/28/2008 rack maybridge local STD ==== ========= ===== === R1A9 HTS04461SC 059 +? R1B9 HTS05414SC 060 +? R1C9 HTS06052SC 061 +? R1D9 HTS06544SC 062 +? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/06/2008 rack maybridge local STD ==== ========= ===== === R1G2 BTB02581SC 007 +? R1G3 BTB12360SC 016 +? R1G7 HTS01756SC 049 +? R1G8 HTS04060SC 057 +? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/11/2008 rack maybridge local STD ==== ========= ===== === R1G4 CD09767 025 - R1G6 GK03165 041 - R1H2 BTB04040 008 - R1H8 HTS04220 058 - 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/12/2008 rack maybridge local STD ==== ========= ===== === R1H3 BTB14130 017 - R1H4 CD10238 026 + R1H5 DP01376 034 + R1H6 GK03187 042 - 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/18/2008 rack maybridge local STD ==== ========= ===== === R1H7 HTS01758 050 - R1A8 HTS02472 051 - R1B8 HTS02489 052 - R1C8 HTS02791 053 - 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/19/2008 rack maybridge local STD ==== ========= ===== === R1D5 CD11549 030 +/-? R1E5 DFP00365 031 +/-? R1F5 DP01247 032 +/-? R1A7 GK03467 043 +/-? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/20/2008 rack maybridge local STD ==== ========= ===== === R1B7 HAN00384 044 +/-? R1C7 HTS00181 045 +/-? R1D7 HTS00266 046 +/-? R1E7 HTS00580 047 +/-? R1F7 HTS01076 048 +/-? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 08/21/2008 rack maybridge local STD ==== ========= ===== === R1D8 HTS03736 054 +/-? R1E8 HTS03891 055 +/-? R1F8 HTS04015 056 +/-? R1A7 GK03467 043 +/-? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 09/04/2008 rack maybridge local STD ==== ========= ===== === R2A2 KM01281 084 +/-? R2A3 KM03273 092 +/-? R2A4 KM07376 101 +/-? R2A5 MWP00086 110 +/-? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 10/20/2008 rack maybridge local STD ==== ========= ===== === R2A6 PD00056 119 R2A7 RH01071 127 R2A8 RJF00759 137 R2A9 S14589 145 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 10/21/2008 rack maybridge local STD ==== ========= ===== === R2A10 SEW00997 153 + R2A11 SEW05555 162 + R2B2 KM01577 085 + R2B3 KM03714 093 ? 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR. # 10/22/2008 rack maybridge local STD ==== ========= ===== === R2B4 KM08121 102 + R2B5 NRB00550 111 + R2B6 PD00768 120 + R2B7 RH01563 128 + 200uL of D2O into each of above, then combine 150 uL of each of the sat'd sol'n, with 60 uL of 500 mM phophate, pH7, 5uL of pHus and 3 uL of TSP (stock 50mM). Used 600 uL to run NMR.