Acid Protease in Nepenthes

Amagase, Shizuko; Nakayama, Shigeru; Tsugita, Akira

doi:10.1093/oxfordjournals.jbchem.a129166

Cited by 32 publications

(6 citation statements)

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“…Bb in S1 File ). This result also demonstrates that the proteases, which cleave the artificial substrate, are stable and active at acidic as well as slightly basic pH ranges, suggesting that these enzymes act like the aspartic proteinases nepenthesin I and II, purified and characterized from several Nepenthes species [ 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 26 ].…”

Section: Resultsmentioning

confidence: 79%

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Nepenthesin Protease Activity Indicates Digestive Fluid Dynamics in Carnivorous Nepenthes Plants

et al. 2015

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Carnivorous plants use different morphological features to attract, trap and digest prey, mainly insects. Plants from the genus Nepenthes possess specialized leaves called pitchers that function as pitfall-traps. These pitchers are filled with a digestive fluid that is generated by the plants themselves. In order to digest caught prey in their pitchers, Nepenthes plants produce various hydrolytic enzymes including aspartic proteases, nepenthesins (Nep). Knowledge about the generation and induction of these proteases is limited. Here, by employing a FRET (fluorescent resonance energy transfer)-based technique that uses a synthetic fluorescent substrate an easy and rapid detection of protease activities in the digestive fluids of various Nepenthes species was feasible. Biochemical studies and the heterologously expressed Nep II from Nepenthes mirabilis proved that the proteolytic activity relied on aspartic proteases, however an acid-mediated auto-activation mechanism was necessary. Employing the FRET-based approach, the induction and dynamics of nepenthesin in the digestive pitcher fluid of various Nepenthes plants could be studied directly with insect (Drosophila melanogaster) prey or plant material. Moreover, we observed that proteolytic activity was induced by the phytohormone jasmonic acid but not by salicylic acid suggesting that jasmonate-dependent signaling pathways are involved in plant carnivory.

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Section: Resultsmentioning

confidence: 79%

“…Proteases in digestive fluid from several species of Nepenthes have also been described early [ 9 ], purified and characterized (e.g. [ 10 , 11 , 12 ]). However, only An et al[ 13 ] cloned nepenthesins from the pitcher tissue of N .…”

Section: Introductionmentioning

confidence: 99%

Nepenthesin Protease Activity Indicates Digestive Fluid Dynamics in Carnivorous Nepenthes Plants

et al. 2015

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“…They have primarily been reported from the pitcher fluid of Nepenthes plants [24,25], and account for the majority of protease activity in the fluid [26]. Purification and characterization of proteases from the pitcher fluid has confirmed that the protease activity was indeed mainly ascribed to nepenthesins [27][28][29]. Subsequently, cDNA clones of nepenthesins were characterized from Nepenthes alata [30] and N. gracilis [31,32].…”

Section: Introductionmentioning

confidence: 99%

Overexpression of Nepenthesin HvNEP-1 in Barley Endosperm Reduces Fusarium Head Blight and Mycotoxin Accumulation

et al. 2020

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Fusarium head blight (FHB) causes substantial losses of yield and quality in grains, both in the field and in post-harvest storage. To date, adequate natural genetic resistance is not available for the control of FHB. This study reports the cloning and overexpression of a barley (Hordeum vulgare L.) antifungal gene, nepenthesin 1 (HvNEP-1), in the endosperm of barley grains. Transgenic barley lines overexpressing HvNEP-1 substantially reduced FHB severity and disease progression after inoculation with Fusarium graminearum or Fusarium culmorum. The transgenic barley also showed reduced accumulation of the mycotoxin deoxynivalenol (DON) in grain, far below the minimum value allowable for food. Semi-field evaluation of four HvNEP-1 transgenic lines revealed substantial reduction of FHB severity and progression as compared with the control H. vulgare cultivar Golden promise (GP) plants. Our study demonstrated the utility of HvNEP-1 for the control of FHB in barley, and possibly other grains such as wheat and maize.

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“…Nepenthesin (Nep) [5,6] is an acid proteinase secreted in the pitcher of Nepenthes species. So far, the acid proteinases from Nepenthes and Drosera species were only partially purified and poorly characterized [7][8][9][10][11][12]. Although they were shown to be members of aspartic proteinases (APs) [9,11], none of the enzymes secreted from carnivorous plants *Address correspondence to this author at the Laboratory of Molecular Biochemistry, School of Life Science, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan; Fax: +81-426-76-7149; E-mail: kenjitak@ls.toyaku.ac.jp…”

Section: Introductionmentioning

confidence: 99%

Nepenthesin, a Unique Member of a Novel Subfamily of Aspartic Proteinases: Enzymatic and Structural Characteristics

Takahashi

Athauda²,

Matsumoto³

et al. 2005

CPPS

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Carnivorous plants are known to secrete acid proteinases to digest prey, mainly insects, for nitrogen uptake. In our recent study, we have purified, for the first time, to homogeneity two acid proteinases, nepenthesin I (Nep I) and nepenthesin II (Nep II) from the pitcher fluid of Nepenthes distillatoria and investigated their enzymatic and structural characteristics. Both enzymes were optimally active at pH approx. 2.6 toward acid-denatured hemoglobin; the specificity of Nep I toward oxidized insulin B chain appears to be similar, but slightly wider than those of other aspartic proteinases (APs). At or below 50 degrees C both enzymes were remarkably stable; especially Nep I was extremely stable over a wide range of pH from 3 to 10 for over 30 days. This suggests an evolutionary adaptation of the enzymes to their specific habitat. We have also cloned the cDNAs and deduced the complete amino acid sequences of the precursors of Nep I and Nep II from the pitcher tissue of Nepenthes gracilis. Although the corresponding mature enzymes are homologous with ordinary pepsin-type APs, both enzymes had a high content of cysteine residues (12 residues per molecule), which are assumed to form six unique disulfide bonds as suggested by computer modeling and are thought to contribute toward the remarkable stability of Neps. Moreover, the amino acid sequence identity of Neps with ordinary APs, including plant vacuolar APs, are remarkably low (approx. 20%), and phylogenetic comparison shows that Neps are distantly related to them to form a novel subfamily of APs with a high content of cysteine residues and a characteristic insertion, named 'the Nep-type AP (NAP)-specific insertion', including a large number of novel, orthologous plant APs emerging in the gene/protein databases.

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Acid Protease in Nepenthes

Cited by 32 publications

References 0 publications

Nepenthesin Protease Activity Indicates Digestive Fluid Dynamics in Carnivorous Nepenthes Plants

Nepenthesin Protease Activity Indicates Digestive Fluid Dynamics in Carnivorous Nepenthes Plants

Overexpression of Nepenthesin HvNEP-1 in Barley Endosperm Reduces Fusarium Head Blight and Mycotoxin Accumulation

Nepenthesin, a Unique Member of a Novel Subfamily of Aspartic Proteinases: Enzymatic and Structural Characteristics

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