Response of the cytoplasmic and membrane proteome of Corynebacterium glutamicum ATCC 13032 to pH changes.
BACKGROUND
C. glutamicum has traditionally been grown in neutral-pH media for amino acid production, but in a previous article we reported that this microorganism is a moderate alkaliphile since it grows optimally at pH 7.0-9.0, as shown in fermentor studies under tightly controlled pH conditions. We determined the best pH values to study differential expression of several genes after acidic or basic pH conditions (pH 6.0 for acidic expression and pH 9.0 for alkaline expression). Thus, it was interesting to perform a detailed analysis of the pH-adaptation response of the proteome of C. glutamicum ATCC 13032 to clarify the circuits involved in stress responses in this bacterium. In this paper we used the above indicated pH conditions, based on transcriptional studies, to confirm that pH adaptation results in significant changes in cytoplasmatic and membrane proteins.


RESULTS
The cytoplasmatic and membrane proteome of Corynebacterium glutamicum ATCC 13032 at different pH conditions (6.0, 7.0 and 9.0) was analyzed by classical 2D-electrophoresis, and by anion exchange chromatography followed by SDS-PAGE (AIEC/SDS-PAGE). A few cytoplasmatic proteins showed differential expression at the three pH values with the classical 2D-technique including a hypothetical protein cg2797, L-2.3-butanediol dehydrogenase (ButA), and catalase (KatA). The AIEC/SDS-PAGE technique revealed several membrane proteins that respond to pH changes, including the succinate dehydrogenase complex (SdhABCD), F0F1-ATP synthase complex subunits b, alpha and delta (AtpF, AtpH and AtpA), the nitrate reductase II alpha subunit (NarG), and a hypothetical secreted/membrane protein cg0752. Induction of the F0F1-ATP synthase complex beta subunit (AtpD) at pH 9.0 was evidenced by Western analysis. By contrast, L-2.3-butanediol dehydrogenase (ButA), an ATPase with chaperone activity, the ATP-binding subunit (ClpC) of an ATP-dependent protease complex, a 7 TMHs hypothetical protein cg0896, a conserved hypothetical protein cg1556, and the dihydrolipoamide acyltransferase SucB, were clearly up-regulated at pH 6.0.


CONCLUSIONS
The observed protein changes explain the effect of the extracellular pH on the growth and physiology of C. glutamicum. Some of the proteins up-regulated at alkaline pH respond also to other stress factors suggesting that they serve to integrate the cell response to different stressing conditions.
Barriuso-Iglesias
Mónica
M
Instituto de Biotecnología de León (INBIOTEC), Parque Científico de León, Av. Real, 1, 24006. León, Spain. mbari@unileon.es
Schluesener
Daniela
D
Barreiro
Carlos
C
Poetsch
Ansgar
A
Martín
Juan F
JF
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D003593:Cytoplasm / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D020543:Proteome / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013312:Stress, Physiological
D001426:Bacterial Proteins; D008565:Membrane Proteins; D020543:Proteome