DnaB helicase dynamics in bacterial DNA replication resolved by single-molecule studies.

In Escherichia coli, the DnaB helicase forms the basis for the assembly of the DNA replication complex. The stability of DnaB at the replication fork is likely important for successful replication initiation and progression. Single-molecule experiments have significantly changed the classical model of highly stable replication machines by showing that components exchange with free molecules from the environment.

Chitosan-citric acid edible coating to control Colletotrichum gloeosporioides and maintain quality parameters of fresh-cut guava.

This study aimed to verify the action of edible chitosan-citric acid (CHI-CA) coating to control Colletotrichum gloeosporioides and maintain quality parameters of fresh-cut guava. Chitosan was obtained from Litopenaeus vannamei shells using high temperature and short exposure times. The minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of CHI-CA against C.

Stability, transdermal penetration, and cutaneous effects of ascorbic acid and its derivatives.

Topically applied antioxidants exert their benefits by offering protection from damaging free radicals and over-the-counter cosmeceuticals incorporating antioxidants are among the most popular anti-aging products available. One potent antioxidant of particular note, vitamin C, has been extensively utilized because it possesses a variety of other cutaneous benefits including photoprotection from UV A & B, neocollagenesis, inhibition of melanogenesis and improvement of a variety of inflammatory skin disorders. However, the instability of this water-soluble vitamin, together with difficulties associated with its topical delivery, has presented issues for the formulation chemist.

Interspecies variation of Kitasatospora recifensis endophytic from yam bean producing thermostable amylases in alternative media.

An endophytic actinomycete isolated from tubers of yam beam (Pachyrhizus erosus L. Urban) was classified as a novel species nominated Kitasatospora recifensis based in phenotypic and genotypic analysis (16S rDNA gene sequence). Monosporic culture using specific ISP2 media revealed three interspecies, which were identified by DNA southern hybridization (Wild strain 13817 W, Aerial Mycelium strain 13817 AM and Vegetative Mycelium strain 13817 VM).

Laser-induced fluorescence spectrum of 3-vinyl-1H-indene.

The laser-induced fluorescence spectrum of 3-vinyl-1H-indene was recorded between 33,000 and 33,800 cm(-1). An origin band was observed at 33,455 cm(-1) along with several low-frequency modes. With the aid of density functional theory and configuration interaction calculations, the electronic transition was assigned as S1 <-- S0 and the short progression in an 80 cm(-1) mode was identified as a vinyl group torsion.

Agronomic effectiveness of biofertilizers with phosphate rock, sulphur and Acidithiobacillus for yam bean grown on a Brazilian tableland acidic soil.

Phosphate rocks have low available P and soluble P fertilizers have been preferably used in plant crop production, although economic and effective P sources are needed. Experiments were carried out on a Brazilian Typic Fragiudult soil with low available P to evaluate the agronomic effectiveness of phosphate rock (PR) compared with soluble phosphate fertilizer. Yam bean (Pachyrhizus erosus) inoculated with rhizobia (strains NFB 747 and NFB 748) or not inoculated was the test crop.

Structure/function studies on a S-adenosyl-L-methionine-dependent uroporphyrinogen III C methyltransferase (SUMT), a key regulatory enzyme of tetrapyrrole biosynthesis.

The crystallographic structure of the Pseudomonas denitrificans S-adenosyl-L-methionine-dependent uroporphyrinogen III methyltransferase (SUMT), which is encoded by the cobA gene, has been solved by molecular replacement to 2.7A resolution. SUMT is a branchpoint enzyme that plays a key role in the biosynthesis of modified tetrapyrroles by controlling flux to compounds such as vitamin B(12) and sirohaem, and catalysing the transformation of uroporphyrinogen III into precorrin-2.

Pyridine-sugar conjugates as potent inhibitors of enzyme-catalysed glycoside hydrolysis.

A series of O- and N-linked pseudo-disaccharides incorporating simple functionalised pyridines were synthesized and demonstrated potent inhibition of the glucoamylase-catalysed reaction.

Functional properties of yam bean (Pachyrhizus erosus) starch.

The study was carried out in order to determine and establish the functional characters of starch extracted from yam bean (Pachyrhizus erosus (L) Urban) compared with cassava starch. Yam bean is a tropical tuber legume easily grown and holds a great potential as a new source of starch. Yam bean starch shows functional properties which are peculiar to those of most starch root crops.

In vitro properties of a recombinant flavonol synthase from Arabidopsis thaliana.

cDNA corresponding to a flavonol synthase gene from Arabidopsis thaliana was cloned and expressed in Escherichia coli. The recombinant protein was purified to near-homogeneity and the catalytic properties of the enzyme were studied in vitro. Together with kaempferol and apigenin the recombinant protein synthesised the (2R,3S)-cis- and (2S,3S)-trans-isomers of dihydrokaempferol from the (2S)- and (2R)-isomers of naringenin, respectively.

Production and characterization of a thermostable glucoamylase from Streptosporangium sp. endophyte of maize leaves.

Thermostable amylolytic enzymes are currently investigated to improve industrial processes of starch degradation. Streptosporangium sp. an endophytic actinomycete isolated from leaves of maize (Zea mays L.

Effect of sulphur inoculated with Thiobacillus on soil salinity and growth of tropical tree legumes.

A greenhouse experiment was carried out with the objective of evaluating the effects of the elementary sulphur inoculated with Thiobacillus, compared with gypsum, in the amendment of a alluvial sodic saline soil from the Brazilian semiarid region, irrigated with saline water and grown with the tropical legumes leucena and mimosa. The treatments consisted of levels of sulphur (0; 300 and 600 kg/ha) and gypsum (1,200 and 2,400 kg/ha), irrigation using different waters containing the salts NaHCO3, MgCl2, CaCl2, NaCl and KCl, with different electrical conductivities (ECs: 0.2.

Production and characterization of a thermostable alpha-amylase from Nocardiopsis sp. endophyte of yam bean.

Thermostable amylolytic enzymes have been currently investigated to improve industrial processes of starch degradation. Studies on production of alpha-amylase by Nocardiopsis sp., an endophytic actinomycete isolated from yam bean (Pachyrhizus erosus L.

Biosynthesis of vitamin B12: the multi-enzyme synthesis of precorrin-4 and factor IV.

Background: In order to study the biosynthesis of vitamin B12, it is necessary to produce various intermediates along the biosynthetic pathway by enzymic methods. Recently, information on the organisation of the biosynthetic pathway has permitted the selection of the set of enzymes needed to biosynthesise any specific identified intermediate. The aim of the present work was to use recombinant enzymes in reconstituted multi-enzyme systems to biosynthesise particular intermediates.

Precise limits of the N-terminal domain of DnaB helicase determined by NMR spectroscopy.

Two separate N-terminal fragments of the 470-amino-acid Escherichia coli DnaB helicase, comprising residues 1-142 and 1-161, were expressed in E. coli. The proteins were extracted in a soluble fraction, purified, and characterised physically.

Biosynthesis of vitamin B12: the preparative multi-enzyme synthesis of precorrin-3A and 20-methylsirohydrochlorin (a 2,7,20-trimethylisobacteriochlorin).

The Bacillus subtilis genes hemB, hemC and hemD, encoding respectively the enzymes porphobilinogen synthase, hydroxymethylbilane synthase and uroporphyrinogen III synthase, have been expressed in Escherichia coli using a single plasmid construct. An enzyme preparation from this source converts 5-aminolaevulinic acid (ALA) preparatively and in high yield into uroporphyrinogen III. The Pseudomonas denitrificans genes cobA and cobI, encoding respectively the enzymes S-adenosyl-L-methionine:uroporphyrinogen III methyltransferase (SUMT) and S-adenosyl-L-methionine:precorrin-2 methyltransferase (SP2MT), were also expressed in E.

Expression, purification and characterisation of the product from the Bacillus subtilis hemD gene, uroporphyrinogen III synthase.

Uroporphyrinogen III synthase, the product of the hemD gene, is the enzyme responsible for the cyclisation of the linear tetrapyrrole, hydroxymethylbilane. The hemD gene isolated from Bacillus subtilis was manipulated by PCR to enable direct cloning behind a synthetic ribosome-binding site downstream of tandem bacteriophage lambda PR and PL promoters in a pCE30-derived vector. Following thermal induction of transcription, the resulting plasmid (pPS21) directed the synthesis of uroporphyrinogen III synthase.

A structural model for the Escherichia coli DnaB helicase based on electron microscopy data.

The DnaB protein is the major replicative DNA helicase in Escherichia coli. It hydrolyzes ATP to promote its translocation in the 5' to 3' direction on single-stranded DNA templates, facilitating the separation of strands of duplex DNA in its path. This places it on the lagging strands at replication forks during chromosomal DNA replication.

Genetics and enzymology of the B12 pathway.

The chemical complexity of vitamin B12 suggests that its formation may involve a large number of enzymic steps. However, until recently, little was known of the number, mechanism and stereochemical course of the many enzymic interconversions that are essential to vitamin B12 biosynthesis. In response to this the French groups led by Francis Blanche and Joel Crouzet have carried out extensive investigations into the genetic and biochemical organization of this remarkable biosynthetic pathway.

The 92-min region of the Escherichia coli chromosome: location and cloning of the ubiA and alr genes.

A cosmid (pND320) bearing 42.5 kb of Escherichia coli chromosomal DNA, including the genes between xylE and ssb near minute 92 on the linkage map, was isolated by selection for complementation of a dnaB mutation. Known nucleotide (nt) sequences were used to align restriction maps in this region to the physical map of the chromosome (coordinates 4319.

Purification and properties of urease from the cyanobacterium Anabaena cylindrica.

A purification procedure has been developed by which urease activity in extracts from the cyanobacterium Anabaena cylindrica was enriched 500-fold. The procedure involves MgSO4 precipitation at 55 degrees C and chromatography on hydroxylapatite and diethylaminoethyl sephadex. Its molecular weight was measured by sedimentation equilibrium in an airfuge to be 197,000 +/- 2000 with an estimated subunit molecular weight of 32,000 as determined by polyacrylamide gel electrophoresis.

Enriched sources of Escherichia coli replication proteins. The dnaG primase is a zinc metalloprotein.

Primase, the product of the Escherichia coli dnaG gene, is the enzyme responsible for RNA primer synthesis on both template strands at replication forks during chromosomal DNA synthesis. The dnaG gene was modified by replacement of the natural ribosome-binding site with one complementary to the 3' end of 16S rRNA, and then inserted downstream of tandem bacteriophage lambda PR and PL promoters in the pUC9-derived vector pCE30. Following thermal induction of transcription, the resulting plasmid pPL195 directed synthesis of primase activity to levels corresponding to approx.

Modified bacteriophage lambda promoter vectors for overproduction of proteins in Escherichia coli.

A new series of expression vectors that direct high-level overproduction of gene products in Escherichia coli is described. All contain strong bacteriophage lambda promoters, PR and PL, arranged in tandem so that both promote transcription into genes inserted into or between unique restriction sites. The vectors also direct expression of the lambda cI857 gene (from its natural promoter, PM), which enables their use in any E.

Care of the elderly has its rewards.

Nursing the elderly is often seen as fit only for those who are unskilled or professionally underdeveloped. This unenlightened view leads to nurses working in this challenging area being looked down on by their colleagues outside the speciality. It is time these attitudes were changed.