The application of two‐dimensional polyacrylamide gel electrophoresis to medical microbiology: Molecular epidemiology of viruses and bacteria

Cash, Phillip

doi:10.1002/elps.1150120721

Cited by 14 publications

(13 citation statements)

References 66 publications

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“…As found for other bacteria, changing the in vitro culture conditions leads to altered protein expression [20] The H. injluenzae genome sequence is an important step towards understanding gene expression in relation to the pathogenesis of this bacterial group. Analyses of H .…”

Section: Analysis Of Haemophilus Proteins By 2-d Pagementioning

confidence: 99%

Development of a Haemophilus two‐dimensional protein database

et al. 1997

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Members of the Haemophilus genus are responsible for various human infections including respiratory infections and meningitis. The complete nucleotide sequence of the Rd strain of Haemophilus influenzae has been reported and represents a valuable resource to investigate gene expression within this bacterial group. We described previously the application of two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) to characterise the proteins of Haemophilus influenzae (Cash et al., Electrophoresis 1995, 16, 135-148). We have extended these data with comparative studies of the proteins from other members of the Haemophilus genus (specifically H. parainfluenzae, H. haemolyticus and H. parahaemolyticus) to identify homologous proteins and, by extension, the genes encoding them, among these bacteria. The proteins extracted from each of these bacterial isolates were compared by coelectrophoresis to the 2-D protein profile of the reference nontypable strain of H. influenzae (HI-64443) used as the basis for the 2-D protein database. A composite reference 2-D protein profile of HI-64443 was derived from three independent analyses of the soluble bacterial proteins. Between 21% and 37% of the HI-64443 proteins from the reference 2-D protein profile comigrated with proteins in the other isolates from the Haemophilus genus. This compared with 62% and 64% comigration when HI-64443 was compared with the Eagan and Rd strains of H. influenzae, respectively. The 2-D protein profile of the Rd strain of H. influenzae was compared to that of HI-64443 by coelectrophoresis; 64% of the proteins detected for the Rd strain comigrated with proteins found for HI-64443 when analysed in parallel. The capacity of 2-D PAGE to investigate global interactions of gene expression was applied to the analysis of superoxide dismutase (SOD) expression in H. influenzae strain Eagan. A "knock-out" mutant in the sodA gene which encodes [Mn]-SOD was characterised with respect to protein synthesis compared to the parental isolate. From these analyses, the primary product of sodA was provisionally identified as a protein with a molecular mass of 25500 Da and an estimated pI of 6.55. Quantitative changes in the expression of two other proteins in the SOD mutant were detected by comparison with the parental isolate. These data are discussed in relation to the development of a 2-D protein database for H. influenzae and related bacteria to investigate genome homologies and gene expression.

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Section: Analysis Of Haemophilus Proteins By 2-d Pagementioning

confidence: 99%

Development of a Haemophilus two‐dimensional protein database

et al. 1997

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“…2-D PAGE can be used to link certain gel patterns to specific diseases or modifications of specific marker proteins, and this technique can also be applied to answer molecular epidemiological questions in medical microbiology (1,4,7,10,11,15,16,19,20,25). The studies mentioned represent some examples of the wide variety of organisms and questions concerning bacterial protein expression that may be analyzed by the 2-D gel technique.…”

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Clustering of Clinical Strains of Helicobacter pylori Analyzed by Two-Dimensional Gel Electrophoresis

Enroth

Åkerlund

Sillén

et al. 2000

Clin Diagn Lab Immunol

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Strain variations of Helicobacter pylori have been tested by numerous methods and compared among different patient groups. The aim of this study was to investigate whether H. pylori expresses disease-specific proteins that can be detected by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). H. pylori strains isolated from duodenal ulcer, gastric cancer, and gastritis patients were analyzed. Extensive variation in spot patterns was observed between the strains, but a dendrogram analysis revealed that some strains within each disease group clustered together. Eight proteins were sequenced and found in the H. pylori genome sequence. 2-D PAGE is a useful method for studies of protein expression and for highlighting the extensive strain variation that H. pylori exhibits.

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“…The origin of the coxsackie B5 virus (CBV5) isolate 3673 (CBV-3673) has been described previously (Cash, 1991;Argo et al, 1992). Two of the virus intracellular proteins, p33 and p39, show mutations which have increased their basic charges and reduced their molecular weights (McLaren et al, 1993).…”

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Genomic and template RNA transcription in a model of persistent enteroviral infection

Jw¹,

Wm²,

Cash³

et al. 1997

J Neurovirol

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Enteroviruses have been implicated in persistent infections of the nervous system and in certain paralytic motor neurone syndromes. Enteroviral persistence may depend on defective transcription, resulting in the abnormal production of equal amounts of genomic and template RNA strands rather than the normal ratio of 60 ± 100 : 1. An in vitro model of a persistent coxsackie virus in human skeletal muscle cells was investigated using in situ hybridisation and a semiquantitative parallel, complementary, reverse transcriptase polymerase chain reaction. The ratio of genomic to template RNA was found to be approximately 60 : 1. We conclude that enteroviral persistence in this in vitro model is not dependent on altered transcription. In vivo, other viral and host factors should be considered.

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The application of two‐dimensional polyacrylamide gel electrophoresis to medical microbiology: Molecular epidemiology of viruses and bacteria

Cited by 14 publications

References 66 publications

Development of a Haemophilus two‐dimensional protein database

Development of a Haemophilus two‐dimensional protein database

Clustering of Clinical Strains of Helicobacter pylori Analyzed by Two-Dimensional Gel Electrophoresis

Genomic and template RNA transcription in a model of persistent enteroviral infection

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