| Autor: |
S. Rocha, W. Verheijen, K. Braeckmans, A. Svendsen, M. Skj∅t, F.C. De Schryver, H. Uji-I, J. Hofkens |
| Rok vydání: |
2007 |
| Předmět: |
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| DOI: |
10.1016/s1574-0641(07)80013-1 |
| Popis: |
In this contribution individual lipase enzymes (PLA1) are investigated in their natural environment, more specific, when released on phospholipid bilayers. After staining the lipid layer, the enzymatic activity could be monitored by the appearance of dark features in the layers by means of wide field microscopy, assuming that the features correspond to reaction products go directly into solution. In this way, real time imaging of the activity is possible. The analysis of the desorbed surface area in the phospholipids bilayer appearing after enzyme injection is then used to estimate the catalytic activity of the enzyme. Variations in speed as a function of time and concentration are observed. Our results show that the enzymatic activity on a natural substrate can be monitored and analysed by the use of wide field microscopy. This approach represents a major step forward in single enzyme studies compared with previous fluorescence-based studies of enzymes where non-natural fluorogenic substrates were used. We also improve the time resolution as compared to AFM measurements on bilayers probing the same enzymatic process. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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