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K Sen

Publications and source records attributed to K Sen.

At least 19 recordsLinked to original sources
PubMed1992

Effects of Bacillus thuringiensis var. israelensis 20-kDa protein on production of the Bti 130-kDa crystal protein in Escherichia coli.

A 20-kDa protein of Bacillus thuringiensis var. israelensis (Bti) has been shown to be necessary for the efficient expression of the 27-kDa mosquitocidal protein gene in Escherichia coli. We have investigated the effects of this 20-kDa protein on the expression of two 130-kDa mosquitocidal protein genes (cryIVA, cryIVB) in E. coli by supplying the 20-kDa protein gene in trans. When a recombinant plasmid, pLH4BX, which was constructed to express cryIVA under the E. coli lac promoter on pUC19, coexisted with the 20-kDa protein gene, a striking increase in production of the fused CryIVA was detected. This was not accompanied by an increase in the amount of intracellular mRNA, suggesting that 20-kDa protein exerts a posttranscriptional effect. We conclude that the 20-kDa protein also stimulates the production of 130-kDa protein in E. coli.

Amino Acid Sequence↗
PubMed1991

Trimerization of an in vitro synthesized OmpF porin of Escherichia coli outer membrane.

The assembly of outer membrane proteins of Escherichia coli was examined using the OmpF porin as a model. Since this protein is made as a precursor, which is processed to a protein of Mr 37,000 before being assembled into trimers in the outer membrane, we synthesized a modified OmpF, which lacked 16 out of 22 amino acid residues from its signal sequence, in a coupled transcription-translation system. This modified protein resembled the unfolded, monomeric OmpF in its electrophoretic behavior, but much of the protein apparently existed in a more tightly folded conformation as it was recognized by a monoclonal antibody specific to a surface epitope of the native, trimeric OmpF porin. At least some conformers of this protein could be further incorporated into outer membrane or lipopolysaccharide bilayers, and assembled into trimers. The trimers formed were trypsin-resistant and heat-stable in sodium dodecyl sulfate up to 70 degrees C, thus showing the characteristics of the native trimeric protein. These results extend our earlier observation that OmpF monomer secreted by spheroplasts of E. coli can be trimerized in vitro (Sen, K., and Nikaido, H (1990) Proc. Natl. Acad. Sci. U.S. A 87, 743-747) and show that the trimerization can occur, albeit at a low efficiency, with porin monomers synthesized in vitro, presumably not contaminated by membrane fragments or other components of the cell envelope. However, comparison of trimerization efficiency of the nascent in vitro product with that of the same product already exposed to aqueous medium, as well as with that of the spheroplast-secreted product, leads us to the working hypothesis that the trimerization process in intact cells is accelerated either by accessory components or by the conformational changes accompanying the secretion through the cytoplasmic membrane and that the reactions observed in this study represent only part of the physiological process.

Amino Acid Sequence↗
PubMed1991

Cloning of a human cDNA expressing a high voltage-activating, TEA-sensitive, type-A K+ channel which maps to chromosome 1 band p21.

Over ten different mammalian genes related to the Drosophila Shaker gene (the Sh gene family) have been identified recently. These genes encode subunits of voltage-dependent K+ channels. The family consists of four subfamilies: ShI genes are homologues of Shaker; ShII, ShIII, and ShIV are homologues of three other Shaker-like genes in Drosophila, Shab, Shaw, and Shal, respectively. We report here the cloning of a human K+ channel ShIII cDNA (HKShIIIC) obtained from a brain stem cDNA library. HKShIIIC transcripts express an atypical voltage-dependent transient (A-type) K+ current in Xenopus oocytes. This current is activated by large membrane depolarizations and is extremely sensitive to the K+ channel blocker TEA unlike most A-type currents. The gene encoding HKShIIIC maps to chromosome 1p21.

Amino Acid Sequence↗
PubMed1991

Lipopolysaccharide structure required for in vitro trimerization of Escherichia coli OmpF porin.

Deep rought mutants, which produce very defective lipopolysaccharides, are unable to export normal levels of porins into the outer membrane. In this study, we showed that lipopolysaccharides from such mutants were also unable to facilitate the trimerization, in vitro, of monomeric OmpF porin secreted by spheroplasts of Escherichia coli B/r. In contrast, lipopolysaccharides containing most or all of the core oligosaccharides were able to facilitate trimerization.

Bacterial Outer Membrane Proteins↗
PubMed1990

Isolation of alpha- and beta-tubulin genes of Plasmodium falciparum using a single oligonucleotide probe.

An oligonucleotide probe (315) specific for the alpha- and beta-tubulin genes of Plasmodium falciparum was synthesized utilizing codon usage of P. falciparum determined from published gene sequences. By screening genomic and cDNA libraries with the oligonucleotide probe, alpha- and beta-tubulin clones were isolated. Positive clones were identified by partial sequencing and comparing the deduced amino acid sequence with the chicken brain alpha- and beta-tubulin amino acid sequences. The beta-tubulin gene was completely sequenced at the genomic level and partially at cDNA level. The deduced polypeptide is 445 amino acids long, shares 88% homology with chicken brain beta-tubulin, and contains two introns of 362 and 163 bp long, respectively. alpha- and beta-tubulin genes of P. falciparum are unlinked and dispersed; more than one copy of each gene may be present. Northern blot analysis of total RNA of the blood-stage parasite indicates the presence of three transcripts of alpha-tubulin (3.3 kb, 2.6 kb, 1.9 kb) and three transcripts of beta-tubulin gene (3.6 kb, 2.9 kb, 2.0 kb). The significance of these transcripts is presently unknown.

Amino Acid Sequence↗
PubMed1990

In vitro trimerization of OmpF porin secreted by spheroplasts of Escherichia coli.

It is not yet clear how bacterial outer membrane proteins reach their correct destination after they are secreted across the cytoplasmic membrane. We show here that porin OmpF is secreted into the medium as a water-soluble monomeric protein by spheroplasts of Escherichia coli. Furthermore, this monomeric porin is taken up by cell envelope preparations or purified lipopolysaccharides in the presence of 0.03% Triton X-100 and is converted correctly into the mature trimeric conformation. These results appear to reproduce a part of the physiological export and targeting steps of this protein.

Bacterial Outer Membrane Proteins↗
PubMed1990

Oligonucleotidase activity of phosphodiesterase from the fruit body of Flammulina velutipes.

A phosphodiesterase (EC 3.1.4.1) was purified to homogeneity from the fruit body of Flammulina velutipes. The enzyme had considerable activity toward oligonucleotides. The Km values were 0.66 mM for ApA, 2.47 mM for (Ap)2A, and 3.03 mM for (Ap)3A. The enzyme hydrolyzed oligodeoxyribonucleotides as well as oligoribonucleotides. The oligoribonucleotides bearing a phosphate residue at the 3' end were not hydrolyzed by the enzyme. The enzyme hydrolyzed the oligoribonucleotides exonucleolytically from the 3' to 5' end. Thus the PDase of F. velutipes is considered to function in vivo as an oligonucleotidase (EC 3.1.13.3), which efficiently converts oligonucleotides to 5'-mononucleotides in the cell.

Chromatography↗
PubMed1988

Porin channels in intact cells of Escherichia coli are not affected by Donnan potentials across the outer membrane.

Donnan potential (interior negative) across the outer membrane of Escherichia coli was measured by the distribution of [14C]choline in a mutant with a deletion through the genes for the active transport of choline. Calculation showed that the presence of membrane-derived oligosaccharides in the periplasm could quantitatively explain the magnitude of the Donnan potential and the periplasmic volume. By measuring the permeability of porin channels in intact cells suspended in solutions of widely different ionic strengths, it was shown that changing Donnan potential from 5 mV to approximately 100 mV had no effect on the permeability of either OmpF or OmpC porin channel toward a zwitterionic compound, cephaloridine. Thus, the "voltage-dependent gating" of porin channel, previously reported from another laboratory, is likely to be an artifact of in vitro reconstitution. The influx of negatively charged compounds, however, was affected by the Donnan potential as expected from the electrolyte diffusion theory.

Algorithms↗
PubMed1987

Studies on orthocephalization: growth behavior of the rat skull in the period 13-49 days as described by the finite element method.

Rat cranial skeletal growth was studied, using a cross-sectional data set, for the period 13-49 days by the application of the concepts of continuum mechanics and the numerical techniques of the finite element method (FEM). In contrast to the methods of conventional craniometry (CM) and roentgenographic cephalometry (RCM) the FEM permits fine scale, reference frame invariant descriptions and analysis of growth behavior. This advantage was demonstrated by a numerical example of the use of FEM. The skull was discretized into a number of two-dimensional, triangular elements, whose enclosed areas corresponded closely to both specific skeletal structures and to related functional matrices. Since it was assumed presently that the growth behavior of all of the points enclosed within a given element was similar, the application of the functional matrix hypothesis permitted an integrated description of the growth of the skeletal structure and functional matrix related to each element. The principal locus of rotation of the facial skull, relative to the cranial base, is the inferior frontoethmoidal articulation, a motion that includes a rigid body rotation. Other active and passive skeletal and visceral growth events associated with orthocephalization were located and described. Finally it was shown that the morphogenetically important growth behavior of other portions of the rat head were not directly involved in orthocephalization.

Aging↗
PubMed1987

Lipid profiles of conidia of Aspergillus niger and a fatty acid auxotroph.

Conidial lipids of the wild-type (V35) Aspergillus niger and its unsaturated fatty acid auxotroph (UFA2) were compared. The wild type contained lower levels (7.6%) of phospholipids and higher levels (28.4%) of glycolipids than the mutant (16.5 and 22.2%, respectively). Oleic (33.4%), linoleic (22.5%), palmitic (12.8%), stearic (7.4%), and linolenic (6.2%) were the main fatty acids of the wild type (V35). The mutant grew only in the presence of unsaturated fatty acid having at least one delta 9cis double bond, and its conidial fatty acid profile was influenced by the exogenous acid. Analyses of the fatty acids of UFA2 grown in the presence of different fatty acid supplements support the original view that the mutant is defective in delta 9-desaturase activity.

Aspergillus niger↗
PubMed1987

Biochemical studies on acetate non-utilizing mutants of Aspergillus terreus IRRL 16043.

The strain Aspergillus terreus IRRL 16043 can utilize glucose as well as acetate as a sole carbon source. Thirty-nine mutants were isolated from the wild-type by treatment with a chemical mutagen, N-methyl-N'-nitro-N-nitrosoguanidine (MNTG) which could not utilize acetate as a sole carbon source, and were designated as acetate non-utilizing (acu). By complementation and biochemical analyses they were divided into three functional groups, acu A, acu B and acu C lacking isocitrate lyase, malate synthase and acetyl-CoA synthetase activity, respectively.

Acetate-CoA Ligase↗
PubMed1987

Effects of pre-weaning undernutrition on 21 day-old male rat skull form as described by the finite element method.

Conventional Roentgenographic Cephalometric Methods (RCM) have certain conceptual and geometric constraints that ensure that their descriptions of cephalic growth and comparisons of cephalic form are reference-frame-dependent; and it is impossible to determine which, if any, RCM reference frame provides a biologically more correct description or comparison. The use of the concepts of continuum mechanics and of the numerical techniques of the Finite Element Method (FEM) overcomes these constraints and provides reference-frame-independent (invariant) growth descriptions and form comparisons. In the FEM, the structure of the head is subdivided (discretized) into a number of smaller, finite, elements. The FEM then describes the growth behavior, or compares the form, of the entire continuum of points enclosed with each element independently; in RCM only the behaviors, or comparisons, of the landmark points are described. The FEM is described and its use illustrated by a comparison of the cephalic forms of 4 groups of weanling (21 d old) rats. The 1st (control) group is derived from normally nourished dams, while the other 3 groups are derived from dams who were malnourished throughout the weaning period in 3 group specific manners. The FEM permitted reference-frame-independent comparisons of the 4 different, and group specific, head forms. These differences were reflected in the forms of each of the finite elements. The differences in element forms between the 4 groups were shown to be related primarily to nutritionally produced alterations of either neurocranial or splanchocranial viscera. These differences were expressed in the alterations of the forms of the continua enclosed within the boundaries of the several finite elements, and secondarily in the form of the cranial skeleton.

Animals↗
PubMed1986

Cytochrome b is necessary for the effective processing of core protein I and the iron-sulfur protein of complex III in the mitochondria.

The effect of cytochrome b on the assembly of the subunits of complex III into the inner mitochondrial membrane has been studied in a mutant of yeast (W-267, Box 6-2) that lacks a spectrally detectable cytochrome b and synthesizes a shortened form of apocytochrome b. We recently reported that several cytochrome b-deficient mutants contained significantly diminished amounts of core proteins I and II as well as the iron-sulfur protein, but contained equal amounts of cytochrome c1 compared to the wild type (K. Sen and D. S. Beattie, Arch. Biochem. Biophys. 242, 393-401, 1985). In the present study, the time course of processing of precursors of both core protein I and the iron-sulfur protein which had accumulated in cells treated with the uncoupler carbonyl m-chlorophenyl hydrazone (CCCP) was noted to be significantly lower in the mutant compared to the wild type. The amounts of the mature forms of these proteins in mitochondria pulse labeled under different conditions was also considerably decreased at all times studied. The synthesis of both proteins appeared to be unaffected in the mutant, as the precursor forms of both proteins accumulated to the same extent when processing in vivo was blocked by CCCP. Furthermore, translation of RNA in a reticulocyte lysate in vitro indicated that the messenger RNAs for both proteins were present in the mutant and translated with equal efficiency. The import into isolated mitochondria of the precursor forms of the iron-sulfur protein synthesized in the cell-free system was also decreased in the mutant mitochondria. In addition, the precursor form was bound to the exterior of the mitochondrial membrane where it was sensitive to digestion with proteases. By contrast, the synthesis and processing of cytochrome c1 appeared to be unaffected in these mutants. These results suggest that cytochrome b is necessary for the proper processing and assembly of both core protein I and the iron-sulfur protein, but not for cytochrome c1, into complex III of the inner mitochondrial membrane.

Biological Transport↗
PubMed1985

Decreased amounts of core proteins I and II and the iron-sulfur protein in mitochondria from yeast lacking cytochrome b but containing cytochrome c1.

The effect of cytochrome b on the assembly of the subunits of complex III into the inner mitochondrial membrane has been studied in four mutants of yeast that lack a spectrally detectable cytochrome b and do not synthesize apocytochrome b. Quantitative analysis of intact mitochondria by immunoprecipitation or immunoblotting techniques with specific antisera revealed that the core proteins and the iron-sulfur protein were decreased 50% or more in the mitochondria from the mutants as compared to the wild type. Sonication of wild-type mitochondria did not result in any decrease in any of these proteins from the membrane; however, sonication of mitochondria from the four mutants resulted in a further decrease in the amount of these proteins suggesting that they are not as tightly bound to the mitochondrial membrane in the absence of cytochrome b. By contrast, the amounts of cytochrome c1 in the mitochondria, as determined both spectroscopically and immunologically, were not significantly affected by the absence of cytochrome b. In addition, no loss of cytochrome c1 was observed after sonication of the mitochondria suggesting that this protein is tightly bound to the membrane. These results suggest that the processing and/or assembly of these subunits of complex III into the mitochondrial membrane is affected by the absence of cytochrome b.

Cytochrome b Group↗
PubMed1985

Finite element method modeling of craniofacial growth.

The application of the concepts of continuum mechanics and of the numerical techniques of the finite element method permits the development of a new and potentially clinically useful method of describing craniofacial skeletal growth. This new method differs from those associated with customary roentgenographic cephalometry in that its descriptions and analyses are invariant; that is, they are independent of any method of registration and superimposition. Such invariance avoids the principal geometric constraint explicit in all analytical methods associated with conventional roentgenographic cephalometry. The conceptual and mathematical bases of the finite element method (FEM) are presented and illustrated by the numerical and graphic descriptions of the two-dimensional growth of the rat skull, for which two sets of longitudinal growth data are used. In practice, the FEM permits analysis of the skull at a scale significantly finer than previously possible, by considering cranial structure as consisting of a relatively large number of contiguous finite elements. For each such element, independently, it is then possible to describe and depict both the magnitude and the direction of temporal size and shape changes occurring in that element relative to itself at some initial time. It is emphasized that such descriptions are completely independent of any local reference frame.

Aging↗
PubMed1985

An unsaturated fatty acid mutant of Aspergillus niger with partially defective delta 9-desaturase.

The wild-type Aspergillus niger (V35) does not require fatty acids for growth. Four unsaturated fatty acid auxotrophs designated as UFA1, UFA2, UFA3, and UFA4 have been produced from this organism by treating the conidia of the wild-type strain with a mutagen, N-methyl-N'-nitro-N-nitrosoguanidine, followed by isolation on media containing monounsaturated fatty acids and the nonionic detergent, Brij 58. Optimal growth of the mutants comparable with that of the wild type was achieved with medium supplemented with C16 or C18 unsaturated fatty acids containing at least one cis double bond at the delta 9 position. Some other fatty acids (18:1 delta 11 cis and 16:1 delta 9 trans) support growth to some extent. The mutants do not grow at all in the presence of saturated fatty acids. Fatty acid analyses of the mutant, UFA2, grown in the presence of different fatty acid supplements reveal that it may be defective in a desaturase system. Experiments with unlabeled and [1-14C]palmitoyl-CoA have shown that the microsomes of the mutant (UFA2) contain a partially defective delta 9-desaturase system.

Aspergillus niger↗
PubMed1985

Lipid profiles of Aspergillus niger and its unsaturated fatty acid auxotroph, UFA2.

A comparative study of the mycelial lipid composition of a wild strain (V35) and one unsaturated fatty acid auxotroph (UFA2) of Aspergillus niger has been performed. The lipid composition of both strains are qualitatively the same but quantitatively different. All the strains contain the following phospholipids: cardiolipin, phosphatidylethanolamine, phosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylcholine, and phosphatidylserine; and triglycerides, diglycerides, monoglycerides, ergosterol, and sterol esters as the neutral lipids; mono- and di-galactosyl diglyceride as the major glycolipids along with small amounts of the corresponding mannose analogs. Phosphatidylethanolamine and phosphatidylcholine constitute the bulk of the phospholipids. The mutant (UFA2) contains a higher level of glycerides and lower levels of sterol (both free and esterified form), phospholipids, and glycolipids than the wild type. Aspergillus niger contains C16 to C18 saturated and unsaturated fatty acids. Small amounts of long-chain (C20 to C24) and short-chain (C10 to C14) saturated and unsaturated acids are also present. Linoleic, oleic, and palmitic are the major acids, stearic and linolenic acids being minor ones. UFA2 grows only in the presence of unsaturated fatty acid (C16 or C18) and accumulates a higher concentration of supplemented acid which influences its fatty acid profile.

Aspergillus niger↗