Biomedical Engineering Reference
In-Depth Information
The proteolytic activity of
Acacia caven
CE was notably enhanced by the addition of
12mM cysteine, but it was completely inactivated by incubation with 0.1 and 10mM of E64,
HgCl
2
and 0.1mM of PMSF after 30 min. This behavior is characteristic of cystein proteases,
although serine proteases can not be discarded before further purification steps were carried
out. Preferences of the
Acacia caven
CE on the p-nitrophenyl esters of different amino acids
were also tested.
Acacia caven
CE showed high activity for Phe, Ala, Gln and Pro derivatives
(unpublished).
In addition of high optimum pH and thermostability, many parameters are involved in
selecting a protease for detergents, such as compatibility with detergent components, e.g.
surfactants, perfumes and bleaches [34, 35].
Acacia caven
proteases (CE) were assayed in the presence of non-ionic and ionic
surfactant at different concentrations and temperatures as were previously described in
Material and methods. Proteolytic activities were all retained after being incubated 30 min
under above conditions. Nevertheless, proteolytic activities were modified when the time of
incubation increased at 60 min (Figure 3). After 1 h at 60° C, the residual activity of
Acacia
caven
CE showed a sharp decreasing with every studied surfactant because of the thermal
inactivation of the proteases. After 1 h of incubation below 40° C, the residual activities of
Acacia caven
CE ranged between 27 and 100 % and the effect of the surfactant concentration
was negligible with the exception of Triton X100. This surfactant was the only one capable of
reducing the proteolytic activity when the temperature and the concentration increased.
In spite of the above results,
Acacia caven
CE was able to retain more than 55 % of initial
activity when it was incubated with six commercial detergents, after 1 h at 37° C (Figure 4).
The observed decreasing on CE activity was attributed to other detergent components than the
detergent own proteases because of these were previously inactivated freezing (24 h) and
overheating (95° C, 1h).
120
100
80
60
40
20
0
1
2
3
4
5
6
7
Figure 4. Effect of different commercial detergent on the proteolityc activity of
Acacia caven
CE, after
1h of incubation at 37º C. (5 mg/ml) Z-Ala p-nitrofenil ester was used as substrate. (1. Woolite, 2.
Ariel, 3. Ace, 4. Skip, 5. Ala Matic, 6. Enzimax, 7. None)
