Use of isoenzyme and protein phenotypes to discriminate between six Pratylenchus species from Great Britain

Six species of Pratylenchus indigenous to Great Britain, P. crenatus, P. fallax, P. neglectus, P. penetrans, P. pinguicaudatus and P. thornei were analysed by slab gel electrophoresis to compare protein patterns and isoenzyme phenotypes of esterase, glucose-6-phosphate dehydrogenase, isocitrate dehydrogenase, phosphoglucose isomerase and phosphoglucomutase. Multiple electromorphs were obtained from all enzymes examined. The results demonstrated that isoenzyme phenotypes are useful to supplement the morphological characterisation of these nematode species. Pair-wise comparisons of the six species were performed giving coefficients of similarity in the range 11–41%. A dendrogram of the six species, generated from the five enzyme banding patterns, gave two groups: group 1 contained P. pinguicaudatus, P. fallax and P. thornei and group 2 contained P. penetrans, P. neglectus and P. crenatus.     

Influence of iron depletion on growth kinetics, siderophore production, and protein expression of Staphylococcus aureus

Abstract Growth rates, siderophore secretion, and bacterial proteins of two clinical isolates of Staphylococcus aureus were studied over 72 h of growth in iron-supplemented and iron-restricted chemically defined media. Under iron restriction the growth rates were decreased to different extents depending on the strain. Production of siderophore was detected in the mid-exponential and stationary phases of growth. The expression of iron-regulated proteins of 81, 23, and 17 kDa was time-dependent, associated with the same stage of growth, and might be involved in siderophore efficiency.     

Exhaustive enumeration of protein conformations using experimental restraints.

We present an efficient new algorithm that enumerates all possible conformations of a protein that satisfy a given set of distance restraints. Rapid growth of all possible self-avoiding conformations on the diamond lattice provides construction of alpha-carbon representations of a protein fold. We investigated the dependence of the number of conformations on pairwise distance restraints for the proteins crambin, pancreatic trypsin inhibitor, and ubiquitin. Knowledge of between one and two contacts per monomer is shown to be sufficient to restrict the number of candidate structures to approximately 1,000 conformations. Pairwise RMS deviations of atomic position comparisons between pairs of these 1,000 structures revealed that these conformations can be grouped into about 25 families of structures. These results suggest a new approach to assessing alternative protein folds given a very limited number of distance restraints. Such restraints are available from several experimental techniques such as NMR, NOESY, energy transfer fluorescence spectroscopy, and crosslinking experiments. This work focuses on exhaustive enumeration of protein structures with emphasis on the possible use of NOESY-determined distance restraints.     

Changes in intracellular proteolysis in Escherichia coli during prolonged growth with a low concentration of dihydrostreptomycin

Abstract Escherichia coli was cultured with a low concentration of dihydrostreptomycin (2.5 μg/ml.). Growth was similar to untreated controls for 10 h after which a slow decline in growth rate occurred; growth ceased after 20 h. Intracellular catabolism of pulse-labelled protein synthesised at various points during the antibiotic treatment increased during the first 10 h, but during the second 10 h proteolysis progressively declined to almost control values. The production of an aberrant proteolytic system is one possible explanation.     

Immunological relationships and a genetic interpretation of major and minor acidic proteins in human parotid saliva

Isoelectric focusing was performed on parotid salivas selected for their electrophoretic phenotypes of proline-rich acidic salivary proteins. Fractions encompassing narrow pH regions were pooled and examined by polyacrylamide gel electrophoresis. Isoelectric focusing yielded partial purification of major and minor acidic proline-rich proteins which were subsequently compared by immunoelectrophoresis and double immunodiffusion against goat anti-human parotid saliva. Cross-reactivity without spurring between all fractions containing major Pr proteins in both immunoelectrophoresis and double immunodiffusion suggests that these proteins are immunologically very similar or identical.This study was supported in part by an award from the American Cancer Society Institutional Grant IN-88F to Fels Research Institute.     

Separation of macromolecules in isotachophoresis systems involving single or multiple counterions

The isotachcophoresis principle provides unique opportunities for rational designs of fractionation procedures involving charged molecules. Theoretically any two charged molecules that are soluble under the experimental conditions involved can be physically separated if their electrophoretic net mobilities differ only slightly in the electrophoresis medium used. A theoretical and practical outline is presented that enables the reader to set up this fractionation system and on a rational basis develop fractionation procedures for a given set of charged macromolecules by isotachophoresis with simple and well characterized ampholytes as spacer substances. The planning of preparative experiments in this approach is based on results obtained from rapid analytical screens on a microgram scale. The report includes an appendix containing the theoretical basis for computation of buffer compositions in the isotachophoretic steady state with mono/polyvalent constituents in systems involving one or more counterions and controlled amounts of interferiong ions.     

Cell free synthesis of some storage protein subunits by polyribosomes and RNA isolated from developing seeds of pea (Pisum sativum L.)

Polyribosomes which have template activity in the wheat germ system have been isolated from developing pea seeds. Some of the translation products have identical mobilities to the vicilin and legumin subunits by SDS-PAGE. Certain products were specifically immunoprecipitated with antisera prepared against purified vicilin and legumin fractions. Various RNA fractions including poly A-rich RNA have also been isolated from polyribosomes and shown to direct the synthesis of polyripeptides whose properties are similar to the storage protein subunits. The results are discussed in relationship to other investigations with seed storage protein biosynthesis in vitro.Abbreviations DTT dithiothreitol - SDS-PAGE SDS-polyacrylamide gel electrophoresis - TCA tricarboxylic acid     

Application of the ‘profiles of relationship’ method to distantly related proteins

Summary An analysis is made of the applicability of the recently published profiles of relationship method for establishing evolutionary relatedness among proteins by using the distantly related proinsulin and neurotoxin protein se-quences as a test object. The method is based on a simultaneous group analysis of both the frequency of acceptance of mutations and their genetic code inter-changeability. Regularities in the patterns of the profiles, which reflect decreased similarity with the passage of time, are established for typical cases of closely related, distantly related and unrelated proteins. This makes it possible to distinguish distantly related from unrelated proteins without extensive statistical randomization procedures. New evidence is stated in favour of a previously suggested definition of interchangeability which does not consider the third base in the codon. The applicability of the profiles of relationship method is examined on the distant relationship between proinsulin and the snake and scorpion neurotoxins which has been established previously by means of conventional approaches.     

Properties of a major α-globulin of rice endosperm

Globulin was isolated from milled rice (Oryza sativa, line IR480-5-9) by 5% NACl extraction and was precipitated by (NH₄)₂SO₂ or by dialysis against water. The extract was purified by repeated isoelectric precipitation at pH 4.5. The major globulin fraction (40%) exhibited one band by electrophoresis at pH 4.5 but two bands at pH 8.3. Similarly, one sharp peak was shown by sedimentation corresponding to 1.41S (α-globulin) in acetic acid (pH 2) and NaOH (pH 11.7) but a broad asymmetric peak was obtained at pH 6.7, 8.3 and 8.9. Gel filtration of the α-globulin at pH 6.5 exhibited 2 proteins with MW 20 000 and 98 000. The results suggest a pH dependent aggregation phenomenon. The two proteins could not be separated by DEAE-cellulose chromatography. SDS-polyacrylamide electrophoresis of α-globulin revealed one subunit with MW 18 000. This α-globulin is poorer in lysine and histidine but richer in cystine, methionine, arginine, tyrosine and glutamic acid than whole milled rice proteinfa]Taken part from the M.S. thesis of AAP from the University of the Philippine at Los Baños (1977).