Growth phase-dependent biosynthesis of Nep, a halolysin-like protease secreted by the alkaliphilic haloarchaeon Natrialba magadii.
OBJECTIVE
The alkaliphilic haloarchaeon Natrialba magadii secretes a halolysin-like protease (Nep) that is active and stable in high salt and in organic solvents, which represents a potential resource for biocatalysis in low water activity conditions. In this study, the effect of the growth stage on Nep biosynthesis was examined.


RESULTS
Nep mRNA and extracellular protease activity were measured by RT-PCR and azocaseinolytic activity determination, respectively. Increased abundance in Nep mRNA was observed in Nab. magadii cells with culture age, which correlated with accumulation of extracellular protease activity. Moreover, a 'stationary phase behavior' on synthesis of Nep was evidenced in low-density cultures incubated with stationary phase medium.


CONCLUSIONS
nep gene expression is up-regulated during the transition to the stationary phase in response to 'factors' (metabolite and/or regulatory molecule) occurring in high-density cultures of Nab. magadii. Although the identity of these molecules remains to be determined, preliminary evidence suggests that they are hydrophobic and stable in high salt and high pH values (3.5 mol l(-1) NaCl, pH 10).


CONCLUSIONS
This study contributes to gain insight into the regulation of haloarchaeal protease biosynthesis, facilitating the large-scale production of this extremozyme for basic studies or potential applications.
Paggi
R A
RA
Instituto de Investigaciones Biológicas, FCEyN-UNMDP, Funes 3250, Mar del Plata, Argentina.
Madrid
E A
EA
D'Alessandro
C P
CP
Cerletti
M
M
De Castro
R E
RE
D019843:Archaeal Proteins / Q000096:biosynthesis* / Q000737:chemistry; D002364:Caseins / Q000378:metabolism; D004795:Enzyme Stability; D020869:Gene Expression Profiling; D019848:Gene Expression Regulation, Archaeal; D006216:Halobacteriaceae / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D010447:Peptide Hydrolases / Q000096:biosynthesis* / Q000737:chemistry; D012331:RNA, Fungal / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012492:Salts
D019843:Archaeal Proteins; D002364:Caseins; D012331:RNA, Fungal; D012333:RNA, Messenger; D012492:Salts; C026018:azocasein; D010447:Peptide Hydrolases