Glutamine synthetase from the marine cyanobacteria Prochlorococcus spp: characterization, phylogeny and response to nutrient limitation.
The regulation of glutamine synthetase (EC 6.3.1.2) from Prochlorococcus was previously shown to exhibit unusual features: it is not upregulated by nitrogen starvation and it is not inactivated by darkness (El Alaoui et al. (2001) Appl Environ Microbiol 67: 2202-2207). These are probably caused by adaptations to oligotrophic environments, as confirmed in this work by the marked decrease in the enzymatic activity when cultures were subjected to iron or phosphorus starvation. In order to further understand the adaptive features of ammonium assimilation in this cyanobacterium, glutamine synthetase was purified from two Prochlorococcus strains: PCC 9511 (high-light adapted) and SS120 (low-light adapted). We obtained approximately 100-fold purified samples of glutamine synthetase electrophoretically homogeneous, with a yield of approximately 30%. The estimated molecular mass of the subunits was roughly the same for both strains: 48.3 kDa. The apparent Km constants for the biosynthetic activity were 0.30 mM for ammonium, 1.29 mM for glutamate and 1.35 mM for ATP; the optimum pH was 8.0. Optimal temperature was surprisingly high (55 degrees C). Phylogenetic analysis of glnA from three Prochlorococcus strains (MED4, MIT9313 and SS120) showed they group closely with marine Synechococcus isolates, in good agreement with other studies based on 16 S RNA sequences. All of our results suggest that the structure and kinetics of glutamine synthetase in Prochlorococcus have not been significantly modified during the evolution within the cyanobacterial radiation, in sharp contrast with its regulatory properties.
El Alaoui
Sabah
S
Departamento de Bioquímica y Biología Molecular, Universidad de Córdoba, Edificio Severo Ochoa, planta 1, Campus de Rabanales, 14071-Córdoba, Spain.
Diez
Jesús
J
Toribio
Fermín
F
Gómez-Baena
Guadalupe
G
Dufresne
Alexis
A
García-Fernández
Jose M
JM
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000145:classification / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D000458:Cyanobacteria / Q000201:enzymology* / Q000235:genetics; D005974:Glutamate-Ammonia Ligase / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D007501:Iron / Q000378:metabolism; D008969:Molecular Sequence Data; D009584:Nitrogen / Q000378:metabolism; D010758:Phosphorus / Q000378:metabolism; D010802:Phylogeny; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D012623:Seawater; D016415:Sequence Alignment
D001426:Bacterial Proteins; D000644:Quaternary Ammonium Compounds; D010758:Phosphorus; D018698:Glutamic Acid; D000255:Adenosine Triphosphate; D007501:Iron; D005974:Glutamate-Ammonia Ligase; D009584:Nitrogen