A sacrificial millipede altruistically protects its swarm using a drone blood enzyme, mandelonitrile oxidase.

Yuko Ishida,Yasumasa Kuwahara,Yayoi Ichiki,Takuya Yamaguchi,Atsutoshi Ina,Masashi Morita,Mohammad Dadashipour,Yasuhisa Asano

doi:10.1038/srep26998

Yuko Ishida, Yasumasa Kuwahara + Show 6 more

Open Access

https://doi.org/10.1038/srep26998

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Abstract

Soldiers of some eusocial insects exhibit an altruistic self-destructive defense behavior in emergency situations when attacked by large enemies. The swarm-forming invasive millipede, Chamberlinius hualienensis, which is not classified as eusocial animal, exudes irritant chemicals such as benzoyl cyanide as a defensive secretion. Although it has been thought that this defensive chemical was converted from mandelonitrile, identification of the biocatalyst has remained unidentified for 40 years. Here, we identify the novel blood enzyme, mandelonitrile oxidase (ChuaMOX), which stoichiometrically catalyzes oxygen consumption and synthesis of benzoyl cyanide and hydrogen peroxide from mandelonitrile. Interestingly the enzymatic activity is suppressed at a blood pH of 7, and the enzyme is segregated by membranes of defensive sacs from mandelonitrile which has a pH of 4.6, the optimum pH for ChuaMOX activity. In addition, strong body muscle contractions are necessary for de novo synthesis of benzoyl cyanide. We propose that, to protect its swarm, the sacrificial millipede also applies a self-destructive defense strategy—the endogenous rupturing of the defensive sacs to mix ChuaMOX and mandelonitrile at an optimum pH. Further study of defensive systems in primitive arthropods will pave the way to elucidate the evolution of altruistic defenses in the animal kingdom.

Highlights

We identify the novel enzyme that catalyzes the synthesis of benzoyl cyanide from mandelonitrile
The purified enzyme was capable of synthesizing benzoyl cyanide from mandelonitrile in vitro (Fig. 1a; see Supplementary Fig. S2), the conditions (0.1 U of ChuaMOX, 1 μmol of mandelonitrile, and 100 μl of aqueous buffer) for which were estimated based on values from an animal extract and crude homogenate[15]
After one hour of incubation at pH 8, 80% of the activity remained between 25 °C and 50 °C, and all activity was lost at 70 °C (Fig. 1f). These results suggest that ChuaMOX is a stable enzyme similar to hydroxynitrile lyase, which was previously isolated from C. hualienensis (ChuaHNL) and considered as a potential industrial biocatalyst for the synthesis of cyanohydrins[8]

Summary

Introduction

We identify the novel enzyme that catalyzes the synthesis of benzoyl cyanide from mandelonitrile. The enzyme is classified as an oxidase. We characterize its enzymatic activity, physico-chemical properties, and localization. The substrate and the enzyme are not colocalized; they separately accumulate in defensive sacs and in blood respectively. Synthesis of benzoyl cyanide likely occurs by, endogenously rupturing the membranes of the defensive sacs using strong body muscle contractions during defensive behavior. We discuss this self-destructive defense of the millipede for protection of its swarm by synthesizing benzoyl cyanide from mandelonitrile through mandelonitrile oxidase

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