The HPLC analysis of peptides before and after the protease activity of O. oeni on NMF from red
wine shows three new peaks, 6 higher and one lower in the chromatograph obtained after the enzyme
action. It produces 148.7 mg l-1 of individual amino acids showing a difference of 109 mg l-1 with respect
to the amino acids produced from white wine. This property is important considering the lower protein
concentration in red wine.
The LAB are able to respond to changes in nitrogen availability by regulating the activity of the
proteolytic system to ensure proper nitrogen balance in the cell. It was found that the synthesis of many
exoproteins is influenced, in part at least, by the levels of individual nutrients in the extracellular
environment. Taking account the stress conditions for O.oeni in wine, the regulation of a strain
exoprotease production and secretion under unfavorable conditions was studied [10]. Starved cells after 2
h incubation at 30°C in citrate buffer 0.05 mmol l-1 pH 5 showed greater extracellular proteolytic activity
than at the onset of starvation. In the presence of 60 mg l-1 SO2 and 8% or 12% ethanol (both
compounds normally present in wine), the proteolytic activity was higher and the role of Ca2+ in
maintaining the attachment of the proteinase to the cell was determined. In starvation condition the
exoprotease production is subject to catabolite repression by glucose and its analogue non-metabolizable
2-DOG. This effect is relieved by the addition of cAMP. The addition of chloramphenicol immediately
stopped protease formation, showing that the appearance of extracellular protease in starvation
conditions resulted from rapid export of the product of de novo protein synthesis, rather than secretion of
protease accumulated intracellulary. The chloramphenicol effect also suggests that the protease activity
was not a result of passive cells lysis. This fact was confirmed by the absence of protease activity in the
supernatant from the antibiotic treated cells showing that it is a result of deliberate release.