Identification, characterization and deduced amino acid sequence of the dominant protease from Kudoa paniformis and K. thyrsites: A unique cytoplasmic cysteine protease

Valerie A. Funk, Robert W. Olafson, Monique Raap, Derek Smith, Laura Aitken, Jody D. Haddow, Diana Wang, Jennifer A. Dawson-Coates, Robert D. Burke, Kristina M. Miller

Research output: Contribution to journalArticlepeer-review

26 Citations (Scopus)

Abstract

Kudoa paniformis and Kudoa thyrsites (Myxozoa: Myxosporea) infections are associated with severe proteolysis of host muscle tissue post-mortem. The present study was undertaken to identify and characterize the protease responsible for myoliquefaction and determine mechanisms controlling protease function in vivo. N-terminal sequence analysis of partially purified protease from hake muscle infected with K. paniformis and K. thyrsites revealed a 23 amino acid sequence that aligned with cysteine proteases. Enzyme inhibition assays confirmed the presence of an essential active site cysteine residue. Using the above K. paniformis amino acid sequence data, a corresponding cDNA sequence from K. thyrsites plasmodia was elucidated revealing a cathepsin L proenzyme (Kth-CL). The translated amino acid sequence lacked a signal sequence characteristic of lysosomal and secreted proteins suggesting a unique cytoplasmic location. Only the proenzyme form of Kth-CL was present in Atlantic salmon muscle anti-mortem but this form became processed in vivo when infected muscle was stored at 4 °C. The proenzyme of Kth-CL showed uninhibited activity at pH 6.0, negligible activity at pH 6.5 and no measurable activity at pH 7.0 whilst the processed protease showed stability and function over a broad pH range (pH 4.5-8.8). The pH dependent processing and function of Kth-CL was consistent with histidine residues in the proregion playing a critical role in the regulation of Kth-CL.

Original languageEnglish
Pages (from-to)477-489
Number of pages13
JournalComparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
Volume149
Issue number3
DOIs
Publication statusPublished - Mar 2008

Keywords

  • Cathepsin L protease
  • K. paniformis
  • Kudoa thyrsites
  • Myoliquefaction
  • Myxosporean
  • Proenzyme
  • pH regulation

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Molecular Biology

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