[Validation of the use of aequorin for cytoplasmic free calcium determination by chemiluminescence in Streptococcus pneumoniae].
In the extracellular pathogen Streptococcus pneumoniae, transformable by soluble DNA, calcium transport is shown to play a key role for vegetative growth, developement of competence for genetic transformation and experimental virulence. To get a more precise localisation of Ca2+ in the cell, we cloned the cDNA of apoaequorine in the chromosome of Streptococcus pneumoniae. This allowed the reconstitution of the acquorine system and chemoluminescence measurements of the cytoplasmic free calcium concentration in the bacteria. Intracellular free Ca2+ is 2 microM at the steady state and can reach 14 microM when calcium is added to the bacterial suspension. Increase in free Ca2+ in response to an imposed Ca2+ gradient depends on the initial velocity (Vi) of the DMB-sensitive Ca2+ transport, showing that changes in cytoplasmic Ca2+ involve active transport.
Chapuy-Regaud
S
S
Laboratoire de Génétique et Physiopathologie de Streptococcus pneumoniae, Pseudomonas aeruginosa et bactéries apparentées EA 3036 CHU Rangueil, Université Paul Sabatier 31403 Toulouse, France.
Jones
H E
HE
Campbell
A K
AK
Trombe
M C
MC
D000331:Aequorin / Q000235:genetics; D001693:Biological Transport, Active; D002118:Calcium / Q000032:analysis* / Q000378:metabolism; D003593:Cytoplasm / Q000737:chemistry; D007202:Indicators and Reagents; D008163:Luminescent Measurements; D011994:Recombinant Proteins; D013296:Streptococcus pneumoniae / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure
D007202:Indicators and Reagents; D011994:Recombinant Proteins; D000331:Aequorin; D002118:Calcium