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A fully modular vector system for the optimization of gene expression in Escherichia coli.

The new expression vector system CYTEXP is designed to facilitate the optimization of both transcription and translation in Escherichia coli, while at the same time allowing the exchange of its major components using unique restriction sites. In vitro mutagenesis can be performed in situ using single-stranded DNA generated from the bacteriophage f1 ORI sequence. The basic vector pCYTEXP1 bears a synthetic copy of the intercistronic sequence that enhances the translation of the E. coli atpE gene. Reading frames can be inserted directly downstream of this sequence. The bacteriophage lambda promoters, the atpE sequence, the bacteriophage fd transcriptional terminator, the f1 ORI, and the amp antibiotic resistance gene are all borne on exchangeable "modules." Thus, both the efficiency and the conditions of expression of cloned genes can be readily optimized.

Antigens, Bacterial↗

Modular organization of motor behavior in the frog's spinal cord.

The complex issue of translating the planning of arm movements into muscle forces is discussed in relation to the recent discovery of structures in the spinal cord. These structures contain circuitry that, when activated, produce precisely balanced contractions in groups of muscles. These synergistic contractions generate forces that direct the limb toward an equilibrium point in space. Remarkably, the force outputs, produced by activating different spinal-cord structures, sum vectorially. This vectorial combination of motor outputs might be a mechanism for producing a vast repertoire of motor behaviors in a simple manner.

Animals↗

Building protein structure and function from modular units.

Many proteins in multicellular organisms are made from combinations of several, clearly identifiable, autonomously folding domains or modules. The structures of many of the constituent modules and some module pairs are now known. This review briefly describes some of the recent X-ray crystallographic and nuclear magnetic resonance (NMR) structural work on modules 'dissected' from proteins that are often large, membrane-bound and glycosylated. These include important proteins involved in cell adhesion, clotting, fibrinolysis and signalling. The structure and function of the intact proteins is discussed in the light of the recent structural work.

Animals↗

MOL3D--a modular and interactive program for molecular modeling and conformational analysis: II. Extended modules.

The new release of MOL3D, a molecular modeling program written in FORTRAN, contains not only enhanced graphic capabilities, but also an improved module for intermolecular calculations that allows rigid and flexible docking. Various interfaces have been added to some well-known and widely diffused programs, such as MM2, AMBER and MOPAC, and to the Cambridge Crystallographic Database. Finally a graph manager and a samples database have been added, which allow efficient searches with various requirements concerning structural templates, pharmacophoric three-dimensional (3D) constraints, and the field of biological activity, if any.

Computer Graphics↗

METIER (Modular Ecotoxicity Tests Incorporating Ecological Relevance) for difficult substances--III. Effects of medium renewal and use of a carrier on the bioavailability of parathion.

In this work we compared the effects of medium renewal and use of a carrier (ethanol) in the lethal toxicity of parathion to Daphnia magna Straus. Actual concentrations measured in test solutions with and without ethanol were not significantly different, indicating that there is no need for a carrier solution to assist dissolution of parathion, up to a concentration of 5 mg litre(-1). LC50 values at 48 h from bioassays with and without medium renewal were identical, indicating that in our experimental conditions, degradation of parathion was not a significant consideration. Similar 48-h LC50 values were obtained in all tests, with or without ethanol present. Thus, potential interactions between carrier and toxicant had no effect on the lethal toxicity of parathion to D. magna.

Journal Article↗

Modular analysis of the control of complex metabolic pathways.

The understanding of the functioning of the intact cell would be simplified appreciably if it were possible first to analyze particular modules of cell physiology separately, and then to integrate the information so as to yield understanding of the control structure in terms of the mutual regulation of the modules. Here we develop a quantitative method based on Metabolic Control Analysis that makes this possible: The relevant properties of the modules are contained in "overall" elasticity coefficients, which reflect the changes in fluxes in the module upon a small variation of the environment of the module, allowing the latter to attain steady state. We show how overall control coefficients, which reflect the control exerted by the processes catalyzed by each module, can be expressed into the overall elasticity coefficients. We derive corresponding summation and connectivity theorems. A number of possible divisions of physiological systems into modules is discussed. This work is a generalization of previous analyses of overall control properties in that it allows for multiple fluxes to connect the modules, and reaction stoichiometries of any complexity.

Cell Physiological Phenomena↗

Modular organization of ON and OFF responses in the cat lateral geniculate nucleus.

Multiunit mapping techniques were used to investigate whether or not units with ON- and OFF-center receptive fields occur in an orderly arrangement in the cat lateral geniculate nucleus. In the A layers, ON- and OFF-center sites were partially segregated in the dimension parallel to the laminae, with ON-center sites predominant at the dorsal surface of each layer and OFF-center sites relatively more numerous at the ventral surface of the layer. Microelectrode tracks were placed to traverse the visual field map in the nucleus parallel to projection lines, parallel to isoelevation lines and parallel to isoazimuth lines. The tracks placed parallel to projection lines contained long sequences of units of the same center sign, providing evidence that ON and OFF zones are organized in vertical modules which span much or all of the thickness of a layer. The tracks placed parallel to isoelevation and isoazimuth lines contained alternating clusters of ON- and OFF-center sites. These clusters were larger in the dimension representing isoelevation than in the dimension representing isoazimuth. Taken together, these results indicate that the cat lateral geniculate nucleus contains ON and OFF modules oriented perpendicular to the layers.

Action Potentials↗

Modular design of the Enterococcus hirae muramidase-2 and Streptococcus faecalis autolysin.

The mature forms of the extracellular muramidase-2 of Enterococcus hirae and Streptococcus faecalis autolysin have very similar primary structures. Each consists of an active-site-containing N-terminal domain fused to a multiple-repeat C-terminal domain. Polypeptide segments occurring at equivalent places in these two bacterial wall lytic enzymes have homologues in two phage lysozymes and in three functionally unrelated proteins, illustrating the principle that protein molecules frequently are constructed from modules that are linked in a single polypeptide chain.

Amino Acid Sequence↗

Organization of the biosynthetic gene cluster for rapamycin in Streptomyces hygroscopicus: analysis of the enzymatic domains in the modular polyketide synthase.

The three giant multifunctional polypeptides of the rapamycin (Rp)-producing polyketide synthase (RAPS1, RAPS2 and RAPS3) have recently been shown to contain 14 separate sets, or modules, of enzyme activities, each module catalysing a specific round of polyketide chain extension. Detailed sequence comparison between these protein modules has allowed further characterisation of aa that may be important in catalysis or specificity. The acyl-carrier protein (ACP), beta-ketoacyl-ACP synthase (KS) and acyltransferase (AT) domains (the core domains) have an extremely high degree of mutual sequence homology. The KS domains in particular are almost perfect repeats over their entire length. Module 14 shows the least homology and is unique in possessing only core domains. The enoyl reductase (ER), beta-ketoacyl-ACP reductase (KR) and dehydratase (DH) domains are present even in certain modules where they are not apparently required. Four DH domains can be recognised as inactive by characteristic deletions in active site sequences, but for two others, and for KR and ER in module 3, the sequence is not distinguishable from that of active counterparts in other modules. The N terminus of RAPS1 contains a novel coenzyme A ligase (CL) domain that activates and attaches the shikimate-derived starter unit, and an ER activity that may modify the starter unit after attachment. The sequence comparison has revealed the surprisingly high sequence similarity between inter-domain 'linker' regions, and also a potential amphipathic helix at the N terminus of each multienzyme subunit which may promote dimerisation into active species.

Amino Acid Sequence↗

Modular automation in PET tracer manufacturing: application of an autosynthesizer to the production of 2-deoxy-2-[18F]fluoro-D-glucose.

A compact autosynthesizer was developed and used successfully for the production of 2-deoxy-2-[18F]fluoro-D-glucose [18FDG] from gaseous acetyl hypo[18F]fluorite. The autosynthesizer performs a sequence of general purpose synthesis procedures named Synthesis Unit Operations (SUO's). Each SUO is controlled through execution of a digital control algorithm with a BASIC language subroutine. This automatic synthesis system is based on two industry standard microcomputer architectures, the IBM PC and STD Bus, and it becomes a component of an evolving distributed microprocessor network of task-dedicated subsystems suitable for automated manufacturing of several useful radiotracers. The yield of 18FDG product using the autosynthesizer and remote manually controlled purification procedures is approximately 20% EOB. Radiochemical purity of this product as measured by thin layer chromatography was 96-99%. Chemical purity of the product was measured to be approximately 96%. 2-Deoxy-2-fluoro-D-mannose impurity from this method was determined to be approximately 4%.

Deoxy Sugars↗