Protein threading by recursive dynamic programming.

We present the recursive dynamic programming (RDP) method for the threading approach to three-dimensional protein structure prediction. RDP is based on the divide-and-conquer paradigm and maps the protein sequence whose backbone structure is to be found (the protein target) onto the known backbone structure of a model protein (the protein template) in a stepwise fashion, a technique that is similar to computing local alignments but utilising different cost functions. We begin by mapping parts of the target onto the template that show statistically significant similarity with the template sequence. After mapping, the template structure is modified in order to account for the mapped target residues. Then significant similarities between the yet unmapped parts of the target and the modified template are searched, and the resulting segments of the target are mapped onto the template. This recursive process of identifying segments in the target to be mapped onto the template and modifying the template is continued until no significant similarities between the remaining parts of target and template are found. Those parts which are left unmapped by the procedure are interpreted as gaps.The RDP method is robust in the sense that different local alignment methods can be used, several alternatives of mapping parts of the target onto the template can be handled and compared in the process, and the cost functions can be dynamically adapted to biological needs.Our computer experiments show that the RDP procedure is efficient and effective. We can thread a typical protein sequence against a database of 887 template domains in about 12 hours even on a low-cost workstation (SUN Ultra 5). In statistical evaluations on databases of known protein structures, RDP significantly outperforms competing methods. RDP has been especially valuable in providing accurate alignments for modeling active sites of proteins.RDP is part of the ToPLign system (GMD Toolbox for protein alignment) and can be accessed via the WWW independently or in concert with other ToPLign tools at http://cartan.gmd.de/ToPLign.html.     

What shapes giant hogweed invasion? Answers from a spatio-temporal model integrating multiscale monitoring data

In this study we simulated the invasion of Heracleum mantegazzianum with a spatiotemporal model that combined a life-cycle matrix model with mechanistic local and corridor dispersal and a stochastic long-distance dispersal in a cellular automaton. The model was applied to the habitat configuration and invader distribution of eight 1?km2 study areas. Comparing the simulations with monitoring data collected over 7?years (2002?C2009) yielded a modelling efficiency of 0.94. We tested the significance of different mechanisms of invasion by omitting or modifying single model components one at a time. Thus we found that the extent of H. mantegazzianum invasion at landscape level depends on both landscape-scale processes and local processes which control recruitment success and population density. Limiting recruitment success (100????30?%) and successionally decreasing the carrying capacity of habitats (max????0) over 30?years significantly improved the projections of the invasion at the landscape level. Local dispersal reached farther than 10?m, i.e. farther than previously assumed, but appeared to be unaffected by wind directions. Long-distance dispersal together with local dispersal dominated the invasion quantitatively. Dispersal through corridors accounted for less invasive spread. Its importance, with respect to invasion speed (number of colonised model grid cells) is probably limited over short periods of time (7?years). Only dispersal along rivers made a significant quantitative contribution to invasion of H. mantegazzianum. We suggest that biotic heterogeneity of suitable habitats is responsible for varying invasion success and that successionally increasing competition leads to declining population densities of H. mantegazzianum over several decades slowing down the spread on the landscape scale.     

Leucyl-leucine methyl ester treatment of donor cells permits establishment of immunocompetent parent----F1 chimeras that are selectively tolerant of host alloantigens

Treatment of murine lymphocytes with L-leucyl-L-leucine methyl ester (Leu-Leu-OMe) selectively removes natural killer cells, cytotoxic T lymphocyte precursors, and the capacity to cause lethal graft-vs-host disease, whereas bone marrow stem cell function and alloantigen-induced L3T4+ T helper function remains intact. The present studies assess the immunocompetence of allogeneic bone marrow chimeras established by reconstituting irradiated (C57BL/6 X DBA/2)F1 (B6D2F1) mice with Leu-Leu-OMe-treated C57BL/6 (B6) bone marrow and spleen cells. Spleen cells from such chimeras were found to have normal B and T cell mitogenic responses. Furthermore, levels of natural-killer cell function were comparable to those observed in B6----B6 syngeneic radiation chimeras established without Leu-Leu-OMe treatment of donor cells. Spleen cells from B6----B6D2F1 mice were identical with B6----B6 or B6 mice in allostimulatory capacity and thus contained no discernible cells of non-H-2b phenotype. Whereas B6----B6D2F1 spleen cells demonstrated alloproliferative and allocytotoxic responses toward H-2k bearing spleen cells, no H-2d specific proliferative or cytotoxic responses could be elicited. B6----B6D2F1 spleen cells did not suppress the generation of anti-H-2d or anti-H-2k proliferative or cytotoxic responses from control B6 spleen cells. Furthermore, addition of rat concanavalin A supernatants did not reconstitute anti-H-2d responses of B6----B6D2F1 chimeric spleen cells. Thus, Leu-Leu-OMe treatment of B6 donor cells not only prevents lethal graft-vs-host disease, but also permits establishment of long-lived parent----F1 chimeras that are selectively tolerant of host H-2 disparate alloantigens, but fully immunocompetent with respect to natural killer cell function, B and T cell mitogenesis, and anti-third party alloresponsiveness.     

Structure of the precursor to an enzyme mediating COOH-terminal amidation in peptide biosynthesis

Many bioactive peptides terminate with an amino acid alpha-amide at their COOH terminus. The enzyme responsible for this essential posttranslational modification is known as peptidyl-glycine alpha-amidating monooxygenase or PAM. We identified cDNAs encoding the enzyme by using antibodies to screen a bovine intermediate pituitary lambda gt11 expression library. Antibodies to a beta-galactosidase/PAM fusion protein removed PAM activity from bovine pituitary homogenates. The 108,207 dalton protein predicted by the complete cDNA is approximately twice the size of purified PAM. An NH2-terminal signal sequence and short propeptide precede the NH2 terminus of purified PAM. The sequences of several PAM cyanogen bromide peptides were localized in the NH2-terminal half of the predicted protein. The cDNA encodes an additional 430 amino acid intragranular domain followed by a putative membrane spanning domain and a hydrophilic cytoplasmic domain. The forms of PAM purified from bovine neurointermediate pituitary may be generated by endoproteolytic cleavage at a subset of the 10 pairs of basic amino acids in the precursor. High levels of PAM mRNA were found in bovine pituitary and cerebral cortex. In corticotropic tumor cells, levels of PAM mRNA and pro-ACTH/endorphin mRNA were regulated in parallel by glucocorticoids and CRF.     

A histomorphometric analysis of trephine biopsies of bone marrow from 65 patients with chronic myeloid leukemia. Classification of patients into subgroups with different survival patterns

A histomorphometric (planimetric) study was performed on trephine biopsies of the bone marrow taken at presentation from 65 patients (31 males and 34 females, with a median age of 48 years) with chronic myeloid leukemia (CML). Specimens from 20 patients (9 males and 11 females, with a median age of 53 years) without any hematologic disorders served as controls. Of the various histologic variables tested, only the counts of neutrophilic granulocytes per 1 sq mm, the ratio of granulocytopoiesis to megakaryopoiesis and the density of reticulin (argyrophilic) fibers revealed a significant correlation with the prognosis. The CML patients were separated into two groups with different survival patterns. Group I (34 patients with a median survival of 24 months) mostly contained cases with the so-called "megakaryocytic subtype" of CML, which is accompanied by variable degrees of fibrosis; group II (31 patients with a median survival of 36 months) mainly contained cases with the "granulocytic subtype," which is not accompanied by myelofibrosis. Among the morphometric parameters, a positive correlation existed between the megakaryocyte count and the reticulin fiber density, which underlines the important role of that cell lineage in fibrillogenesis. There were multiple interrelationships between the histomorphometric variables and the laboratory data. Consequently, multivariate regression methods (using Cox's proportional hazards model) were applied to assess the relative predictive value of the patient characteristics for survival. The derived prognostic model divided the patients into two risk groups, with median survivals of 14 and 41 months, respectively. In order of their entry into the regression model, these variables were percentage of neutrophils in the differential blood count, amount of granulopoiesis, liver size, percentage of peripheral myeloblasts and density of reticulin fibers in the bone marrow. In comparing the two patient groups, based on bone marrow histomorphometric parameters, this model revealed that two of those factors (amount of granulopoiesis and density of reticulin fibers) had a significant correlation with the prognosis.     

Enzymatic dehalogenation of 4-chlorobenzoate by 4-chlorobenzoate dehalogenase from Pseudomonas sp. CBS3 in organic solvents

Summary 4-Chlorobenzoate dehalogenase from Pseudomonas sp. CBS3 showed dehalogenating activity in various organic solvents. In alcohols like methanol (150%) or ethanol (120%) higher activities than in water (100%) were obtained. In apolar solvents like petroleum ether (5%) and nhexane (5%) only trace activities were observed. The solvents did not increase the stability of the enzyme. 4-Chlorobenzoic acid methylester, a substance not soluble in water, was not dehalogenated in organic solvents.     

Leu-Leu-OMe sensitivity of human activated killer cells: delineation of a distinct class of cytotoxic T lymphocytes capable of lysing tumor targets

Sensitivity to L-leucyl-L-leucine methyl ester (Leu-Leu-OMe) was used to characterize the phenotype of human activated killer cells. Natural killer cells (NK) and the precursors of both the alloantigen-specific cytotoxic T lymphocytes (CTL) and the NK-like activated killer cells generated after stimulation with allogeneic cells were deleted from human peripheral blood lymphocytes by preincubation with Leu-Leu-OMe. It was noted, however, that cytotoxic lymphocytes could be generated from Leu-Leu-OMe-treated lymphocyte precursors after 2 to 6 days of culture with the nonspecific mitogen, phytohemagglutinin (PHA). The characteristics of these killer cells indicated that they were a unique population that could be distinguished from other cytotoxic cells. Killing by these cells exhibited slow kinetics in that 18 hr cytotoxicity assays were required to detect full cytotoxic potential. When 18 hr assays were used, PHA-stimulated cytotoxic cells generated from Leu-Leu-OMe-treated lymphocytes were able to kill both NK-sensitive K562 cells and the relatively NK-resistant renal cell carcinoma cell line, Cur. These cytotoxic lymphocytes were HNK-1, Leu-11b (CD16), and OKM1 (CR3)-negative at both the precursor and effector stage of activation. Furthermore, these cells were derived from a CD3-positive precursor. Finally, killing by activated effectors was inhibited by OKT3. Unlike activation of Leu-Leu-OMe-sensitive large granular lymphocytes, generation of these cytotoxic T cells was totally prevented by treatment with mitomycin c before stimulation. Thus, a unique class of tumoricidal T cells can be characterized by resistance of lymphocyte precursors to a concentration of Leu-Leu-OMe, which has been shown to ablate NK, mixed lymphocyte culture-activated NK-like cytotoxic precursors, and the precursors of alloantigen-specific CTL.