Alkaline phosphatase from venom of the endoparasitoid wasp, Pteromalus puparum

Jia-Ying Zhu1,2a, Gong Yin Ye1b*, Qi Fang1c, Cui Hu1d

1State Key Laboratory of Rice Biology & Key laboratory of Molecular Biology of Crop Pathology and Insects of Ministry of Agriculture, Institute of Insect Sciences, Zhejiang University, Hangzhou 310029, China
2Key Laboratory of Forest Disaster Warning and Control of Yunnan Province & Key laboratory of Southwest Mountain Forest Resources Conservation and Utilization of Ministry of Education, Southwest Forestry University, Kunming 650224, China

Abstract

Using chromogenic substrates 5-bromo-4-chloro-3'-indolyl phosphate and nitro blue tetrazolium, alkaline phosphatase (ALPase) was histochemically detected in the venom apparatus of an endoparasitoid wasp, Pteromalus puparum L. (Hymenoptera: Pteromalidae). Ultrastructural observations demonstrated its presence in the secretory vesicles and nuclei of the venom gland secretory cells. Using p-nitrophenyl phosphate as substrate to measure enzyme activity, the venom ALPase was found to be temperature dependent with bivalent cation effects. The full-length cDNA sequence of ALPase was amplified from the cDNA library of the venom apparatus of P. puparum, providing the first molecular characterization of ALPase in the venom of a parasitoid wasp. The cDNA consisted of 2645 bp with a 1623 bp open reading frame coding for 541 deduced amino acids with a predicted molecular mass of 59.83 kDa and pI of 6.98. Using multiple sequence alignment, the deduced amino acid sequence shared high identity to its counterparts from other insects. A signal peptide and a long conserved ALPase gene family signature sequence were observed. The amino acid sequence of this venom protein was characterized with different potential glycosylation, myristoylation, phosphorylation sites and metal ligand sites. The transcript of the ALPase gene was detected by RT-PCR in the venom apparatus with development related expression after adult wasp emergence, suggesting a possible correlation with the oviposition process.

Keywords: histochemistry, cloning

Correspondence: ajiayingzhu001@yahoo.com.cn, b*chu@zju.edu.cn, cgiggs0574@sina.com, dgyye@zju.edu *Corresponding author

Received: 18 June 2008 | Accepted: 18 November 2008 | Published: 4 March 2010

ISSN: 1536-2442 | Volume 10, Number 14

Zhu JY, Ye G Y, Fang Q, Hu C. 2010. Alkaline phosphatase from venom of the endoparasitoid wasp, Pteromalus puparum 15pp. Journal of Insect Science 10:14, available online: insectscience.org/10.14


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