Phenoloxidase Activity of Helix aspersa Maxima (Garden Snail, Gastropod) Hemocyanin

Abstract

The oxygen-transporting protein, hemocyanin (Hc), of the garden snail Helix aspersa maxima (HaH) was isolated and kinetically characterized. Kinetic parameters of the reaction of catalytic oxidation of catechol to quinone, catalyzed by native HaH were determined: the V max value amounted to 22 nmol min(-1) mg(-1), k cat to 1.1 min(-1). Data were compared to those reported for other molluscan Hcs and phenoloxidases (POs). The o-diphenoloxidase activity of the native HaH is about five times higher than the activity determined for the Hcs of the terrestrial snail Helix pomatia and of the marine snail Rapana thomasiana (k cat values of 0.22 and 0.25 min(-1), respectively). The K m values obtained for molluscan Hcs from different species are comparable to those for true POs, but the low catalytic efficiency of Hcs is probably related to inaccessibility of the active sites to potential substrates. Upon treatment of HaH with subtilisin DY, the enzyme activity against substrate catechol was considerably increased. The relatively high proteolytically induced o-diPO activity of HaH allowed using it for preparation of a biosensor for detection of catechol.

Differential Protein Expression in the Hemolymph of Bithynia siamensis goniomphalos Infected with Opisthorchis viverrini

“…The function HMC in invertebrate is not only as a respiratory protein via oxygen transport, but it has also been strongly associated with different immunological processes [60–66]. HMC has been shown to participate in multiple roles of the immune response including phenoloxidase activity [61–64], antiviral [65] and antimicrobial activity [66]. In snails, a study on the transcriptional profile of B .…”

Inhibitory mechanisms of glabridin on tyrosinase

“…Tyrosinase plays a crucial role in melanin biosynthesis pathway that is responsible for melanogenesis of animals [3]. And it actually catalyzes the hydroxylation of tyrosine to L-DOPA (L-3, 4-dihydroxyphenylalanine) and the subsequent oxidation of L-DOPA to DOPA quinone [4,5] which transforms into melanin and other polyphenolic compounds after a series of spontaneous chemical reactions [6]. Overproduction of melanin may lead to a variety of skin diseases, such as melasma, age spots and freckles [7,8].…”

Structural insights into the interaction between molluscan hemocyanins and phenolic substrates: An in silico study using docking and molecular dynamics

“…Prior studies have reported different versions of tyrosinase-based biosensors for detection of phenols in environmental samples [33][34][35][36]. Two recent reports, one from our group [7] and the other by Raynova et al [5] described hemocyanin-based optical biosensors built by immobilizing molluscan hemocyanin on a thin film of chitosan coated on a glass slide. This first-generation sensor was able to detect catechol at concentrations of 5 ppm to 90 ppm.…”

“…However, it suffers from lower affinity for bulky phenolic substrates. Currently, optimal phenoloxidase activity in hemocyanins is obtained by either carrying out limited proteolysis [5] or pretreatment with SDS [7], prior to its immobilization on sensors. Thus, it is important to understand the structural changes caused by the action of proteolytic agents or SDS resulting in an increase in the affinity of the protein for bulky phenolic substrates.…”

“…Recent in vitro studies have detected phenoloxidase activity associated with hemocyanin suggesting its possible role in innate defence mechanisms involving the production of microbicidal pigment, melanin [4]. This activation of enzymatic properties is mediated in vitro by various agents that include proteolytic cleavage by trypsin and chymotrypsin and structural disruption by denaturants like sodium dodecyl sulphate (SDS) [4][5][6][7].…”