12 August 1998 Christian This is the README for the second part of T79, which is the sequence after the suspected HTH (Leu30 - Lys49). The top hits from t79-2nd-sum98.rdb: fssp rep %IDE t79-2nd 1san -8.030 2hddA 50 t79-2nd 1ahdP -7.930 2hddA 50 These four are the same t79-2nd 2hoa -7.930 2hddA 50 homeodomain protein t79-2nd 1hom -7.620 2hddA 50 t79-2nd 1ftz -6.490 2hddA 43 t79-2nd 1wbr -5.770 Immunoglobulin fold t79-2nd 1lea -5.760 1lea 100 same LexA repressor protein t79-2nd 1leb -5.760 1lea 100 t79-2nd 1nedA -5.020 1ned 100 hydrolase t79-2nd 1nedB -5.020 1ned 99 t79-2nd 1nedC -5.020 1ned 100 t79-2nd 2fokA -5.000 2fokA 100 restriction endonuclease t79-2nd 2fokB -5.000 2fokB 100 t79-2nd 1mylA -4.960 1bazA 92 arc repressor t79-2nd 1mylB -4.960 t79-2nd 1mylC -4.960 t79-2nd 1mylD -4.960 t79-2nd 1mylE -4.960 t79-2nd 1mylF -4.960 t79-2nd 1fokA -4.850 2fokA 90 restriction endonuclease t79-2nd 1a1fA -4.700 1aayA 97 zinc finger DNA binding t79-2nd 1a1gA -4.700 1aayA 97 By functionality, those from the 2hddA, 1lea, and 1bazA FSSP families bear looking into. The 1aayA zinc finger is not, as t79 doesn't contain any cysteines (that are required in the motif). 1lea contains beta structure, but the prediction for t79 is all alpha helix. 1bazA uses antiparallel beta strands in the major groove. It also exists as a dimer, but T79 probably doesn't exist as a dimer (PMID:8825776, PMID:8483417) So this leaves 1san and 1ftz. 1san is the top hit for t79-2nd. T79 is a prokaryotic protein, but 1san is a homeodomain protein. While the homeodomain is only known to exist in eukaryotic transcription regulators, 1san has been found to bear similarity to prokaryotic repressors. From PMID:2572329: The structure of the Antennapedia homeodomain from Drosophila melanogaster was determined by nuclear magnetic resonance spectroscopy in solution. It includes three well-defined helices (residues 10-21, 28-38, and 42-52) and a more flexible fourth helix (residues 53-59). **Residues 30-50 form a helix-turn-helix motif virtually identical to those observed in various prokaryotic repressors.** Further comparisons of the homeodomain with prokaryotic repressors showed that there are also significant differences in the molecular architectures. Overall, these studies support the view that the third helix of the homeodomain may function as the DNA recognition site. The elongation of the third helix by the fourth helix is a structured element that so far appears to be unique to the Antennapedia homeodomain. I have created a couple of promising alignments to 2hddA. 2hddA-t79-2nd-cbarrett.hand1.a2m: Despite slightly lower sequence identity, could be the better of the two alignments because of the conservation pattern. 2hddA-t79-2nd-cbarrett.hand1.a2m: This is an full length alignment of t79-2nd to 1hddA. There is one insertion which adds one full turn from the helix. 1ftz still needs to be looked at. Here is the abstract of the paper (PMID:7909851) describing its structure: The three-dimensional structure of a recombinant 70-residue polypeptide containing the complete fushi tarazu (ftz) homeodomain from Drosophila melanogaster has been determined by nuclear magnetic resonance (NMR) spectroscopy in solution. On the basis of 915 upper distance constraints derived from nuclear Overhauser effects and 178 dihedral angle constraints, a group of 20 conformers representing the solution structure of the ftz homeodomain was computed with the program DIANA and energy-minimized with the program OPAL. The average of the pairwise root-mean-square deviations of the individual NMR conformers relative to the mean coordinates is 0.50 A for the backbone atoms N, C alpha and C' of residues 8 to 53. The molecular architecture includes three helices comprising the residues 10 to 21, 28 to 38, and 42 to 52, a loop of residues 22 to 27 between the helices I and II, and a turn of residues 39 to 41 linking the helices II and III. Comparisons with the structure of the mutant Antennapedia homeodomain with Cys39 replaced by Ser, Antp (C39S), shows that the two proteins contain the same molecular fold for residues 8 to 53, whereas the more flexible fourth helix comprising residues 53 to 59 in the Antp (C39S) homeodomain has no counterpart in the ftz homeodomain. Considering that important intermolecular interactions in the DNA complexes with the Antp, engrailed and Mat alpha 2 homeodomains involve the fourth helix, it was rather unexpected that the stability of the complex of ftz with the BS2 operator site was found to be comparable to or even somewhat higher than that of the Antp complex with BS2. Another difference is that the Antp homeodomain is more stable with respect to thermal denaturation, with denaturation temperatures at pH 4.8 of 27 degrees C and 48 degrees C, respectively, for ftz and Antp. Note that 1san/1ftz also have the HTH DNA binding motif, giving T79 two HTH motifs. This is satisfying because -- as far as I have read -- nearly all HTH motifs act in tandem as transcriptional regulators. 13 August 1998 Kevin Karplus Christian seems to have been using a different version of fssp that the 3-5-98 one I've been using. In the one I've been using, 1san and 1ftz are represented by 1fjlA. (Hmm, it looks like pcd/fssp points to an older fssp, I'll update it to point to the one we are now using.) 17 August 1998 Christian I copied the new Makefile from t79 and remade joints. Nothing surprisingly new. 1ftz/t79-2nd-1ftz-global.pw.dist:1ftz 70 -7.26 -7.49 1lea/t79-2nd-1lea-global.pw.dist:1lea 72 -7.20 -9.77 1san/t79-2nd-1san-global.pw.dist:1san 62 -8.94 -9.53 2hddA/2hddA-t79-2nd-global.pw.dist:t79-2nd 80 -3.82 -3.53 My best alignment yet for the second part of t79 is t79-2nd-1san-global.cbarrett1.a2m. I don't know if I'll have time to look for anything better.