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Comparative proteome analysis of subcellular fractions from Borrelia burgdorferi by NEPHGE and IPG
Author(s) -
Nowalk Andrew J.,
Nolder Christi,
Clifton Dawn R.,
Carroll James A.
Publication year - 2006
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200500187
Subject(s) - proteome , borrelia burgdorferi , groel , biology , proteomics , membrane protein , cell fractionation , biochemistry , immobilized ph gradient , membrane , isoelectric focusing , escherichia coli , enzyme , gene , antibody , immunology
Borrelia burgdorferi , the cause of Lyme disease, produces excessive amounts of membrane lipoproteins such as outer surface protein A (OspA) when grown in vitro , and consequently many low or moderately abundant proteins are underrepresented when cell lysates are examined by 2‐DE. We analyzed the B. burgdorferi B31 proteome computationally and by IPG or modified NEPHGE after subcellular fractionation into membrane‐associated and soluble proteins. The B. burgdorferi B31 theoretical proteome is comprised of 1623 proteins and has a mean p I of 8.36 and a median p I of 9.03 with 68% of the proteome possessing a p I ≥7.5. Separation of soluble proteins by IPG resulted in 205 individual spots and identification of 78 protein spots by MALDI‐TOF MS. Separation by modified NEPHGE routinely resulted in approximately 185 soluble and 160 membrane protein spots with the identification of 88 individual protein spots combined by MALDI‐TOF MS. Homologues to GroEL and aminopeptidase I were present in greater amounts in the membrane faction, with enolase at nearly equivalent amounts in the soluble and membrane fractions. Identification of proteins isolated and separated by such methods will enable future determination of proteome changes in membrane and soluble protein fractions as spirochetes adapt to their changing environments.