Comparative zone electrophoresis of catalase of Staphylococcus species isolated from mammalian skin.

Vertical polyacrylamide slab gel electrophoresis was conducted for the catalase enzymes of representative strains of 18 proposed species and subspecies of the genus Staphylococcus. The catalase bands which resulted were predominantly monomorphic within each of the species and differences in catalase mobilities were observed between many of the species. The electrophoretic mobilities of the catalases were supportive to the scheme of classification used. Many strains of certain species demonstrated multiple catalase bands which are suggestive of multimolecular forms of the enzyme. Horizontal starch gel electrophoresis of representative strains of S. capitis produced catalase bands with relative mobilities that were different from those obtained with polyacrylamide electrophoresis, presumably due to a difference in molecular sieving between the gels.     

Electrophoretic and immunological charactorization of rat neurophysin

1. The electrophoretic properties of rat posterior pituitary proteins have been compared on starch gel with those of bovine and porcine neurophysins. 2. [(35)S]-Cysteine was injected into the supraoptic nucleus of male rats and 16-24h later the distribution of labelled neural-lobe protein in starch and polyacrylamide gels was determined. In both systems a single major protein component was found to contain more than 80% of the total recovered radioactivity. Between 5 and 10% of the radioactivity was found in a minor component in polyacrylamide gel. 3. In agar, microimmuno-diffusion and -electrophoresis of the rat neural-lobe proteins gave a single arc with neurophysin antiserum, and after starch-gel electrophoresis this arc was shown to be due to the major labelled component. 4. The molecular weights of the rat neural-lobe proteins were estimated by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. The molecular weight of the major labelled component was found to be 12000. 5. It is concluded that the rat neurophysin consists of one major and possibly one minor component.     

Two previously undetected variants of glutamic-pyruvic transaminase found by acidic polyacrylamide gel electrophoresis.

Two new electrophoretic variants of glutamic-pyruvic transaminase (GPT) have been found by polyacrylamide gel electrophoresis at acidic pH. They appeared to represent a single allele, GPT 2, by the standard method of starch gel electrophoresis. Studies in families show that they are inherited as codominant alleles at the GPT locus. Population frequencies are about the same as those of other rare GPT variants. Their behavior on gels is consistent with both of them having substitutions of histidines in place of uncharged amino acids.     

Enzyme specificity: “pseudopolymorphism” of lactate dehydrogenase in Drosophila melanogaster

An electrophoretic variant in the LDH (l-lactate:NAD oxidoreductase, E.C.1.1.1.27) of Drosophila melanogaster was observed on starch (or polyacrylamide) gels. This variant was found to exhibit an identical isozymic pattern (three isozymes with a decreasing staining density) on starch gel and map position as the Adh locus. On the other hand, anodal polyacrylamide gel electrophoresis in crude extracts has shown LDH to consist of nine bands and ADH of four bands. We have shown that ADH (Alcohol:NAD oxidoreductase, E.C.1.1.1.1) also oxidizes l(+)-lactate or d(–)-lactate with the NAD, while LDH oxidizes ethanol. By using various genetic and biochemical techniques, we have shown that the observed Ldh electrophoretic variant was not a real one and could be attributed to the presence of ADH. We have called this phenomenon pseudopolymorphism, and the problem of enzyme specificity has been examined. The appearance of a band in an assay using lactic acid as a substrate is not sufficient evidence for the presence of LDH. Hence, caution is called for before characterizing an electrophoretic band on a gel as being equivalent to the presence of a genetic locus. Out of the nine electrophoretic zones of activity observed on polyacrylamide gel (or out of the six previously observed) using crude extract, only two (one major and one minor) belong to LDH, as revealed by purified enzyme preparations. Furthermore, purified LDH exhibits activity in two bands on starch gel (out of three observed in crude extracts), which appear in different positions as compared with those of ADH. Finally, one band which responds to the presence of d(–)-lactate but not to l(+)-lactate has been revealed.     

A two-dimensional gel electrophoretic procedure for the separation of complex mixtures of 4–12S RNAs

A two-dimensional gel electrophoretic method suitable for the separation of complex mixtures of RNA species in the size range of 4 to 12 S is described. A 3.6–11% polyacrylamide gradient gel containing a gradient of 0–7 m urea was used in the first dimension, and a transverse 3.6–22.6% polyacrylamide gradient gel containing 5 m urea was used in the second dimension. The method was applied to the separation of total cytoplasmic RNAs from a cellular slime mold. In this method reproducible fingerprints were obtained by the use of visible-marker RNA.     

Identification of 30-kDa fat body protein of Sarcophaga peregrina larvae selectively phosphorylated in the presence of 20-hydroxyecdysone as ribosomal protein S6

Previously, we showed that 20-hydroxyecdysone induces selective phosphorylation of a fat body protein of Sarcophaga peregrina with a molecular mass of about 30,000 (30-kDa protein) (Itoh, K., Ueno, K., & Natori, S. (1985) Biochem. J. 227, 683-688). This paper describes the identification of this 30-kDa protein. From the electrophoretic profile of 40S ribosomal proteins on two-dimensional polyacrylamide gel electrophoresis, the 30-kDa protein was identified as S6.     

苦参和苦豆子的过氧化物酶同工酶谱分析和比较

采用聚丙烯酰胺凝胶电泳技术分析和比较了5个不同产地的苦参和1种苦豆子的过氧化物酶同工酶活性。结果表明：不同产地苦参品种和苦豆子的过氧化物酶同工酶谱迁移率在0.14～0.28之间,共6条酶谱带。聚类分析表明,苦参和苦豆子的亲缘关系较远,而不同产地的苦参种质也存在一定的差异,可分为3类：甘肃成县栽培的苦参与甘肃岷县栽培的苦参为一类,河南卢氏野生苦参单独为一类,河北承德野生苦参和甘肃成县野生苦参为一类。     

Purification and characterization of nucleoside diphosphate kinase from spinach leaves.

Two types of nucleoside diphosphate kinase (NDP kinase I and NDP kinase II) have been purified from spinach leaves to electrophoretic homogeneity. The enzymes were copurified with apparent [35S]GTP-gamma S-binding activities. NDP kinase I, which was not adsorbed to a hydroxyapatite column, and NDP kinase II, which was adsorbed, had molecular weights of 16,000 and 18,000, respectively, as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The molecular weights determined by gel filtration were 92,000 and 110,000, respectively, suggesting that both enzymes are composed of six identical subunits. Minor differences in some amino acids between NDP kinase I and NDP kinase II were observed when both enzymes were analyzed for amino acid composition. The apparent [35S]GTP gamma S-binding activity of purified NDP kinase I and NDP kinase II was found to be due to the formation of a [35S]thiophosphorylated enzyme, which is the intermediate of the NDP kinase reaction.     

Disc electrophoretic studies of hemoglobin dissociation by p-hydroxymercury benzoate

The reaction of excess p-mercurybenzoate with human hemoglobin produces five electrophoretic species on polyacrylamide gel. Only two of these bands have been previously observed when starch gel was employed. The chemical and electrophoretic studies presented in this paper illustrate that the appearance of an “extra” band in the β zone is due to the ability of PAGE to separate the βb₂^PMB ? β^PMB equilibrium to its discrete components. The remaining bands are accounted for by the superior resolution of PAGE over starch gel electrophoresis which allowed the detection of two (αβ)^PMB dimer species.     

The search for hidden enzymatic variation in the aphid Macrosiphum rosae (L.)

Summary Using two different buffer systems and up to 6 different electrophoretic polyacrylamide gel concentrations, hidden enzymatic variability was investigated in samples of the rose aphid Macrosiphum rosae. MDH1, PGM1, SDH, EST and LAP were found to be polymorphic. No additional variation was observed by changing the test conditions compared to those of earlier investigations using starch gel electrophoresis.     

E. Coli Transfer RNA: Fractionation By Electrophoresis on Polyacrylamide Gel

1. The tRNA (4S) of E. coli has been fractionated by electrophoresis on polyacrylamide gel and separated into two electrophoretic classes designated 4S RNA A and 4S RNA B.     

Polyacrylamide Gel Electrophoresis of Corynebacterium diphtheriae: a Possible Epidemiological Aid

In this preliminary study, the use of polyacrylamide gel electrophoresis as an aid in the characterization of Corynebacterium diphtheriae was evaluated and a standardized method was developed. The electrophoretic patterns of 17 gravis, 14 mitis, and 2 intermedius types of C. diphtheriae were compared with the electrophoretic patterns of 5 Robinson and Peeney stock gravis serotype strains. Each of the 5 stock serotype strains had different electrophoretic patterns, although some common bands were present. The 17 gravis strains isolated in the United States showed patterns identical to those of the stock gravis serotype II strain. The 14 mitis strains examined produced 6 different electrophoretic patterns, irrespective of geographical location. One mitis pattern corresponded with the pattern of gravis serological type II. The two intermedius strains examined had identical electrophoretic patterns that resembled the pattern of gravis serotype IV. Polyacrylamide gel electrophoresis of C. diphtheriae strains may prove to be a useful epidemiologic tool in establishing the distribution and occurrence of various C. diphtheriae types.     

High molecular weight ribosomal ribonucleic acid from vegetative hyphae and spores of Streptomyces griseus.

High molecular weight ribosomal ribonucleic acids (rRNAs) were isolated from young vegetative cells and spores of a streptomycin non-producing Streptomyces griseus, and their electrophoretic mobility was compared to each other and to that of rRNAs of Escherichia coli K-12. The electrophoretic mobility of 23 and 16S rRNAs from vegetative cells and spores of S. griseus was identical, but the 23S rRNAs of streptomyces ribosomes migrated more slowly on polyacrylamide gel than those of E. coli ribosomes. Intact, electrophoretically homogenous rRNAs could be isolated from S. griseus (No. 45-H) only in the presence of diethyl 1 pyrocarbonate (DEP), and intact rRNAs could be obtained from spores only if DEP had been added before breaking the spores. Otherwise instead of two distinct bands, three were obtained on polyacrylamide gel.     

Two-Dimensional Protein Patterns in Heterodera glycines

Two-dimensional polyacrylamide gel electrophoretic protein patterns of H. glycines from southern Indiana (Posey County) and northern Indiana (Pulaski County) were largely similar, but many differences existed. The pattern of the Posey isolate was similar to patterns from isolates collected in other areas of the United States. Unique dense protein spots in the pattern of an isolate from Hokkaido, Japan, distinguished it from patterns of six U.S. isolates.     

Serum and tissue alkaline phosphatases in pigs

The alkaline phosphatases from serum, liver, bone and intestine of pigs were separated by starch and polyacrylamide gel electrophoresis. Treatments with neuraminidase, urea, heat, L-homoarginine and L-phenylalanine were performed. Variants of serum alkaline phosphatases were derived from different tissues and hence must be under the control of at least two different loci. Within the intestinal phosphatases, polymorphic electrophoretic patterns were observed among 195 animals.     

Highly purified agarose as stacking gel in sodium dodecyl sulphate/polyacrylamide-gel electrophoresis.

A modified sodium dodecyl sulphage/polyacrylamide-gel-electrophoretic method is described that utilizes highly purified agarose as stacking gel. The same electrophoretic resolution of different marker proteins is found as when polyacrylamide is used as stacking gel, but the background staining seen when polyacrylamide is used as stacking gel is decreased.     

Serum and tissue alkaline phosphatases in pigs*

The alkaline phosphatases from serum, liver, bone and intestine of pigs were separated by starch and polyacrylamide gel electrophoresis. Treatments with neuraminidase, urea, heat, L-homoarginine and L-phenylalanine were performed. Variants of serum alkaline phosphatases were derived from different tissues and hence must be under the control of at least two different loci. Within the intestinal phosphatases, polymorphic electrophoretic patterns were observed among 195 animals.     

MITOCHONDRIAL AND CYTOPLASMIC RIBOSOMES FROM TETRAHYMENA PYRIFORMIS : Correlative Analysis by Gel Electrophoresis and Electron Microscopy

Mitochondrial and cytoplasmic ribosomes from Tetrahymena pyriformis have been isolated and studied by the techniques of polyacrylamide gel electrophoresis and electron microscopy used in conjunction. Although the two ribosome types show the same coefficient of sedimentation (80S) in sucrose gradients, they can be distinguished by gel electrophoresis: mitoribosomes migrate in a single band, considerably slower than the cytoribosome band. Electron microscope observations of negatively stained cytoribosomes show typical rounded or triangular profiles, about 275 x 230 Å; mitoribosome profiles are much larger and clearly elongate, about 370 x 240 Å. An electron-opaque spot delimits two nearly equal size subunits. In mixtures of mito- and cytoribosomes, each type can be recognized by its characteristic electrophoretic mobility and by its distinctive fine structure. Cytoribosomal 60S and 40S subunits each produce a distinct electrophoretic band. On the contrary, neither electrophoretic analysis, using a variety of conditions, nor electron microscopy is able to discern two different subunit types in the single 55S mitoribosomal subunit peak. Electrophoretic analysis of RNA shows that both ribosomal RNA species are present in the mitoribosomal subunit fraction. These results establish that mitoribosomes from T. pyriformis dissociate into two subunits endowed with the same sedimentation coefficient, the same electrophoretic mobility, and a similar morphology.     

A rapid and effecient method for the isolation of proteins from polyacrylamide cells.

A procedure is described for the rapid and efficient electrophoretic elution of protein from polyacrylamide gels which is then collected in a dialysis bag tied to the end of a tube containing the gel slices. To illustrate the method a heterogeneous preparation of alkaline phosphatase was used from which a single homogeneous component was isolated in six hours with a recovery of 86%. The eluted protein is collected in a volume which can easily be kept below 1.5 ml, thus eliminating the need for subsequent concentration. The method has also been used successfully in two other systems in which a human lung tumor-associated antigen and glycogen synthetase from yeast were isolated. Since the method utilizes a standard analytical gel electrophoresis apparatus with no modifications or accessories, it should be immediately applicable for the isolation of many different proteins from polyacrylamide gels.     

A Rapid and Efficient Method for the Isolation of Proteins from Polyacrylamide Gels

A procedure is described for the rapid and efficient electrophoretic elution of protein from polyacrylamide gels which is then collected in a dialysis bag tied to the end of a tube containing the gel slices. To illustrate the method a heterogeneous preparation of alkaline phosphatase was used from which a single homogeneous component was isolated in six hours with a recovery of 86%. The eluted protein is collected in a volume which can easily be kept below 1.5 ml, thus eliminating the need for subsequent concentration. The method has also been used successfully in two other systems in which a human lung tumor-associated antigen and glycogen synthetase from yeast were isolated. Since the method utilizes a standard analytical gel electrophoresis apparatus with no modifications or accessories, it should be immediately applicable for the isolation of many different proteins from polyacrylamide gels.