Fenoloxidáza hmyzu - jak ovlivnit činnost enzymu?
| Title in English | Insect phenoloxidase - How to influence enzymatic activity? |
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| Authors | |
| Year of publication | 2009 |
| Type | Conference abstract |
| MU Faculty or unit | |
| Citation | |
| Description | Phenoloxidase (PO) is the central component of the phenoloxidase cascade. This cascade includes, in addition to the PO enzyme itself, molecules recognizing antigenic compounds that are characteristic for potential pathogens and several serine proteases that mediate transfer of signal about antigen presence and subsequent activation of PO. PO activation is iniciated by cleavage of precursor molecule, which is called prophenoloxidase (pPO) and is synthesized by hemocytes. Insect PO takes part in many immune and non-immune processes such as detection and aggregation of bacteria, nodulation, encapsulation, sclerotization and pigmentation of the cuticle, clot formation and wound healing. PO activity is affected especially by synthesis of pPO, during its activation and subsequently by regulation of activated PO. Using spectrophotometrical measurement of PO activity in larvae and pupae of Galleria mellonella we proved that the PO activity changes during development both in hemolymph and cuticle. pPO synthesis is also influenced by disruption of arachidonic acid metabolism using eicosanoid biosynthesis inhibitors (EBI). Indomethacin causes conspicuous decrease both in PO and pPO activity. On the other hand, high concentrations of phenidone have probably positive effect on PO activity. During in vitro tests factors influencing pPO activation play the key role. The most important are temperature and incubation time. Spontaneous activation of pPO occurs during 5 min at room temperature, but can be drawn away by temperatures under 0C. Total PO levels, that are detectable after chemical activation of overall pPO by methanol, are not influenced neither by temperature nor by incubation time. Action of activated enzyme consists especially in availability and type of substrate. The most suitable substrates for PO are diphenols such as DOPA or hydroquinone. Hydroquinone, in contrast to DOPA, is not bound to solid tissues and therefore can be used for detection of PO activity in cuticle. Furthermore, inhibitors such as phenylthiourea can bind to the active site of the enzyme and completely block PO activity. This work was supported by grant of the GAČR no. 206/09/P470. |
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