Structural diversity of neuronal calcium sensor proteins and insights for activation of retinal guanylyl cyclase by GCAP1.

Neuronal calcium sensor (NCS) proteins, a sub-branch of the calmodulin superfamily, are expressed in the brain and retina where they transduce calcium signals and are genetically linked to degenerative diseases. The amino acid sequences of NCS proteins are highly conserved but their physiological functions are quite different. Retinal recoverin controls Ca(2) (+)-dependent inactivation of light-excited rhodopsin during phototransduction, guanylyl cyclase activating proteins 1 and 2 (GCAP1 and GCAP2) promote Ca(2) (+)-dependent activation of retinal guanylyl cyclases, and neuronal frequenin (NCS-1) modulates synaptic activity and neuronal secretion. Here we review the molecular structures of myristoylated forms of NCS-1, recoverin, and GCAP1 that all look very different, suggesting that the attached myristoyl group helps to refold these highly homologous proteins into different three-dimensional folds. Ca(2) (+)-binding to both recoverin and NCS-1 cause large protein conformational changes that ejects the covalently attached myristoyl group into the solvent exterior and promotes membrane targeting (Ca(2) (+)-myristoyl switch). The GCAP proteins undergo much smaller Ca(2) (+)-induced conformational changes and do not possess a Ca(2) (+)-myristoyl switch. Recent structures of GCAP1 in both its activator and Ca(2) (+)-bound inhibitory states will be discussed to understand structural determinants that control their Ca(2) (+)-dependent activation of retinal guanylyl cyclases

Structural basis for sequence specific DNA binding and protein dimerization of HOXA13.

The homeobox gene (HOXA13) codes for a transcription factor protein that binds to AT-rich DNA sequences and controls expression of genes during embryonic morphogenesis. Here we present the NMR structure of HOXA13 homeodomain (A13DBD) bound to an 11-mer DNA duplex. A13DBD forms a dimer that binds to DNA with a dissociation constant of 7.5 nM. The A13DBD/DNA complex has a molar mass of 35 kDa consistent with two molecules of DNA bound at both ends of the A13DBD dimer. A13DBD contains an N-terminal arm (residues 324 - 329) that binds in the DNA minor groove, and a C-terminal helix (residues 362 - 382) that contacts the ATAA nucleotide sequence in the major groove. The N370 side-chain forms hydrogen bonds with the purine base of A5* (base paired with T5). Side-chain methyl groups of V373 form hydrophobic contacts with the pyrimidine methyl groups of T5, T6* and T7*, responsible for recognition of TAA in the DNA core. I366 makes similar methyl contacts with T3* and T4*. Mutants (I366A, N370A and V373G) all have decreased DNA binding and transcriptional activity. Exposed protein residues (R337, K343, and F344) make intermolecular contacts at the protein dimer interface. The mutation F344A weakens protein dimerization and lowers transcriptional activity by 76%. We conclude that the non-conserved residue, V373 is critical for structurally recognizing TAA in the major groove, and that HOXA13 dimerization is required to activate transcription of target genes

Independent Orbiter Assessment (IOA): Assessment of the electrical power generation/power reactant storage and distribution subsystem FMEA/CIL

The results of the Independent Orbiter Assessment (IOA) of the Failure Modes and Effects Analysis (FMEA) and Critical Items List (CIL) is presented. The IOA effort first completed an analysis of the Electrical Power Generation/Power Reactant Storage and Distribution (EPG/PRSD) subsystem hardware, generating draft failure modes and potential critical items. To preserve independence, this analysis was accomplished without reliance upon the results contained within the NASA FMEA/CIL documentation. The IOA results were then compared to the NASA FMEA/CIL baselines with proposed Post 51-L updates included. A resolution of each discrepancy from the comparison is provided through additional analysis as required. The results of that comparison are documented for the Orbiter EPG/PRSD hardware. The comparison produced agreement on all but 27 FMEAs and 9 CIL items. The discrepancy between the number of IOA findings and NASA FMEAs can be partially explained by the different approaches used by IOA and NASA to group failure modes together to form one FMEA. Also, several IOA items represented inner tank components and ground operations failure modes which were not in the NASA baseline

Wind-tunnel Investigation of Two Airfoils with 25-percent-chord Gwinn and Plain Flaps

Aerodynamic force tests of an NACA 23018 airfoil with a Gwinn flap having a chord 25 percent of the overall chord and of an NACA 23015 airfoil with a plain flap having a 25-percent chord were conducted to determine the relative merits of the Gwinn and the plain flaps. The tests indicated that, based on speed-range ratios, the plain flap was more effective than the Gwinn flap. At small flap deflections, the plain flap had lower drag coefficients at lift-coefficient values less than 0.70. For lift coefficients greater than 0.70, however, the Gwinn flap at all downward flap deflections had the lower drag coefficients