Two-level QSAR network (2L-QSAR) for peptide inhibitor design based on amino acid properties and sequence positions.

In the design of peptide inhibitors the huge possible variety of the peptide sequences is of high concern. In collaboration with the fast accumulation of the peptide experimental data and database, a statistical method is suggested for peptide inhibitor design. In the two-level peptide prediction network (2L-QSAR) one level is the physicochemical properties of amino acids and the other level is the peptide sequence position. The activity contributions of amino acids are the functions of physicochemical properties and the sequence positions. In the prediction equation two weight coefficient sets {ak} and {bl} are assigned to the physicochemical properties and to the sequence positions, respectively. After the two coefficient sets are optimized based on the experimental data of known peptide inhibitors using the iterative double least square (IDLS) procedure, the coefficients are used to evaluate the bioactivities of new designed peptide inhibitors. The two-level prediction network can be applied to the peptide inhibitor design that may aim for different target proteins, or different positions of a protein. A notable advantage of the two-level statistical algorithm is that there is no need for host protein structural information. It may also provide useful insight into the amino acid properties and the roles of sequence positions.

Virtual screening for finding natural inhibitor against cathepsin-L for SARS therapy.

Recently Simmons et al. reported a new mechanism for SARS virus entry into target cells, where MDL28170 was identified as an efficient inhibitor of CTSL-meditated substrate cleavage with IC(50) of 2.5 nmol/l. Based on the molecule fingerprint searching method, 11 natural molecules were found in the Traditional Chinese Medicines Database (TCMD). Molecular simulation indicates that the MOL376 (a compound derived from a Chinese medicine herb with the therapeutic efficacy on the human body such as relieving cough, removing the phlegm, and relieving asthma) has not only the highest binding energy with the receptor but also the good match in geometric conformation. It was observed through docking studies that the van der Waals interactions made substantial contributions to the affinity, and that the receptor active pocket was too large for MDL21870 but more suitable for MOL736. Accordingly, MOL736 might possibly become a promising lead compound for CTSL inhibition for SARS therapy.

Anti-SARS drug screening by molecular docking.

Starting from a collection of 1386 druggable compounds obtained from the 3D pharmacophore search, we performed a similarity search to narrow down the scope of docking studies. The template molecule is KZ7088 (Chou et al., 2003, Biochem Biophys Res Commun 308: 148-151). The MDL MACCS keys were used to fingerprint the molecules. The Tanimoto coefficient is taken as the metric to compare fingerprints. If the similarity threshold was 0.8, a set of 50 unique hits and 103 conformers were retrieved as a result of similarity search. The AutoDock 3.011 was used to carry out molecular docking of 50 ligands to their macromolecular protein receptors. Three compounds, i.e., C(28)H(34)O(4)N(7)Cl, C(21)H(36)O(5)N(6), and C(21)H(36)O(5)N(6), were found that may be promising candidates for further investigation. The main feature shared by these three potential inhibitors as well as the information of the involved side chains of SARS Cov Mpro may provide useful insights for the development of potent inhibitors against SARS enzyme.

Inhibition of PYK2-induced actin cytoskeleton reorganization, PYK2 autophosphorylation and focal adhesion targeting by FAK.

Focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (PYK2) are structurally related tyrosine kinases. They are implicated in regulating actin cytoskeleton organization, a process critical for cell migration, mitosis and tumor metastasis. In this paper, we demonstrate that, although both PYK2 and FAK were expressed and colocalized at focal adhesions in fibroblasts, microinjection of PYK2, but not FAK, in Swiss 3T3 fibroblastic cells led to reorganization of focal adhesions and cell rounding. PYK2-mediated actin cytoskeleton reorganization required the PYK2 N terminus, the focal adhesion targeting (FAT) domain, catalytic activity and autophosphorylation. Remarkably, FAK suppressed PYK2-mediated reorganization of focal adhesions and cell rounding. In addition, FAK inhibited PYK2 autophosphorylation and focal adhesion targeting, which might contribute to FAK-mediated suppression of PYK2's phenotypes. Further analyses demonstrated that the inhibition of PYK2 autophosphorylation required the FAK N terminus but not FAK tyrosine phosphorylation. The FAK FAT domain seemed to be critical for FAK-mediated suppression of PYK2 focal adhesion targeting. Taken together, these results demonstrate that FAK could inhibit PYK2 autophosphorylation, focal adhesion targeting and actin cytoskeleton reorganization, suggesting that the balance between FAK and PYK2 tyrosine kinases is important for regulating cellular morphology, cell migration and cell growth.

Regulation of CDC42 GTPase by proline-rich tyrosine kinase 2 interacting with PSGAP, a novel pleckstrin homology and Src homology 3 domain containing rhoGAP protein.

Proline-rich tyrosine kinase 2 (PYK2), a tyrosine kinase structurally related to focal adhesion kinase (FAK), is implicated in regulating cytoskeletal organization. However, mechanisms by which PYK2 participates in and regulates cytoskeletal organization remain largely unknown. Here we report identification of PSGAP, a novel protein that interacts with PYK2 and FAK and contains multiple domains including a pleckstrin homology domain, a rhoGTPase-activating protein domain, and a Src homology 3 domain. PYK2 interacts with PSGAP Src homology 3 domain via the carboxyl-terminal proline-rich sequence. PSGAP is able to increase GTPase activity of CDC42 and RhoA in vitro and in vivo. Remarkably, PYK2, but not FAK, can activate CDC42 via inhibition of PSGAP-mediated GTP hydrolysis of CDC42. Moreover, PSGAP is localized at cell periphery in fibroblasts in a pleckstrin homology domain-dependent manner. Over expression of PSGAP in fibroblasts results in reorganization of cytoskeletal structures and changes of cellular morphology, which requires rhoGTPase-activating activity. Taken together, our results suggest that PSGAP is a signaling protein essential for PYK2 regulation of cytoskeletal organization via Rho family GTPases.

Regulation of neuregulin signaling by PSD-95 interacting with ErbB4 at CNS synapses.

Neuregulins (NRGs) and their receptors, the ErbB protein tyrosine kinases, are essential for neuronal development, but their functions in the adult CNS are unknown. We report that ErbB4 is enriched in the postsynaptic density (PSD) and associates with PSD-95. Heterologous expression of PSD-95 enhanced NRG activation of ErbB4 and MAP kinase. Conversely, inhibiting expression of PSD-95 in neurons attenuated NRG-mediated activation of MAP kinase. PSD-95 formed a ternary complex with two molecules of ErbB4, suggesting that PSD-95 facilitates ErbB4 dimerization. Finally, NRG suppressed induction of long-term potentiation in the hippocampal CA1 region without affecting basal synaptic transmission. Thus, NRG signaling may be synaptic and regulated by PSD-95. A role of NRG signaling in the adult CNS may be modulation of synaptic plasticity.