Superimposing the 27 crystal protein/inhibitor complexes of ß-secretase to calculate the binding affinities by the linear interaction energy method.

BACE-1 is an important target for designing therapeutic agents for the treatment of Alzheimer's disease. An improved linear interaction energy (LIE) model has been developed to calculate the binding free energies of ß-secretase (BACE-1) by superimposing the 27 crystal BACE-1/inhibitor complexes to put a diverse set of 27 co-crystallized ligands into the binding pocket. These co-crystallized conformations of ligands were set as the initial binding conformations for LIE simulation. The effects of two protein conformations (i.e., 1W51 and 1FKN), two sampling methods (i.e., energy minimization and hybrid Monte Carlo [HMC]), and energy terms were studied. Using 1W51 crystal structure and HMC sampling technique, the best binding affinity model for the full set of ligands was found to have a root-mean-square error of 0.996 kcal/mol.

A binary matrix for improved detection of phosphopeptides in matrix-assisted laser desorption/ionization mass spectrometry.

Application of matrix-assisted laser-desorption/ionization mass spectrometry (MALDI MS) to analysis and characterization of phosphopeptides in peptide mixtures may have a limitation, because of the lower ionizing efficiency of phosphopeptides than nonphosphorylated peptides in MALDI MS. In this work, a binary matrix that consists of two conventional matrices of 3-hydroxypicolinic acid (3-HPA) and alpha-cyano-4-hydroxycinnamic acid (CCA) was tested for phosphopeptide analysis. 3-HPA and CCA were found to be hot matrices, and 3-HPA not as good as CCA and 2,5-dihydroxybenzoic acid (DHB) for peptide analysis. However, the presence of 3-HPA in the CCA solution with a volume ratio of 1:1 could significantly enhance ion signals for phosphopeptides in both positive-ion and negative-ion detection modes compared with the use of pure CCA or DHB, the most common phosphopeptide matrices. Higher signal intensities of phosphopeptides could be obtained with lower laser power using the binary matrix. Neutral loss of the phosphate group (-80 Da) and phosphoric acid (-98 Da) from the phosphorylated-residue-containing peptide ions with the binary matrix was decreased compared with CCA alone. In addition, since the crystal shape prepared with the binary matrix was more homogeneous than that prepared with DHB, searching for 'sweet' spots can be avoided. The sensitivity to detect singly or doubly phosphorylated peptides in peptide mixtures was higher than that obtained with pure CCA and as good as that obtained using DHB. We also used the binary matrix to detect the in-solution tryptic digest of the crude casein extracted from commercially available low fat milk sample, and found six phosphopeptides to match the digestion products of casein, based on mass-to-charge values and LIFT TOF-TOF spectra.