Gene structure, activity and localization of a catalase from intracellular bacteria in Onchocerca volvulus

• Published in: Molecular and biochemical parasitology Vol. 96; no. 1; pp. 69 - 81
• Main Authors: Henkle-Dührsen, Kimberly, Eckelt, Volker H.O, Wildenburg, Gabriele, Blaxter, Mark, Walter, Rolf D
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 30-10-1998
• Subjects: 16S rRNA; Amino Acid Sequence; Animals; Catalase; Catalase - analysis; Catalase - chemistry; Catalase - genetics; Catalase - metabolism; Female; Genes, Bacterial; Genes, rRNA; Immunoenzyme Techniques; Immunolocalization; Intracellular bacteria; Male; Microfilariae - enzymology; Microfilariae - microbiology; Microscopy, Electron; Molecular Sequence Data; Onchocerca volvulus; Onchocerca volvulus - enzymology; Onchocerca volvulus - growth & development; Onchocerca volvulus - microbiology; Onchocerca volvulus - ultrastructure; Open Reading Frames; Phylogenetic analysis; Phylogeny; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Rickettsiaceae - enzymology; Rickettsiaceae - genetics; Rickettsiaceae - ultrastructure; RNA, Bacterial - genetics; RNA, Ribosomal, 16S - genetics; Sequence Alignment; Wolbachia; Immunolocalization; Phylogenetic analysis; 16S rRNA; Catalase; Wolbachia; CAT, catalase; Onchocerca volvulus; Intracellular bacteria
• ISSN: 0166-6851; 1872-9428
• DOI: 10.1016/S0166-6851(98)00109-1

Within the context of studies on the antioxidant enzymes in
Onchocerca volvulus, DNA clones encoding catalase (CAT) were isolated from an
O.
volvulus adult
λzapII cDNA library. Analysis of their nucleotide and encoded amino acid sequences revealed that they derive from intracellular bacteria, rather than the
O.
volvulus nuclear genome. The endobacterial CAT gene was found to lie in a gene cluster, followed by a ferritin gene and an excinuclease gene. The endobacterial CAT gene encodes a functional enzyme capable of detoxifying H
2O
2, demonstrated by producing an active recombinant protein in an
E. coli expression system. The purified 54 kDa protein has CAT activity over a broad pH range, with a specific activity of 103 000±3000 U mg
−1. The optical spectrum of the endobacterial CAT shows that it is a ferric haem-containing protein with a Soret band at 405 nm. To investigate the phylogeny of the intracellular bacterium in
O.
volvulus, a segment of the 16S rRNA gene was amplified from total genomic DNA by a polymerase chain reaction using universal eubacterial primers. A phylogenetic analysis of the
O. volvulus-derived 16S rRNA sequence revealed that the endobacterium belongs to a distinct
Wolbachia clade of the order Rickettsiales. Onchocercomata and biopsies containing different onchocercal species were immunohistochemically stained using polyclonal antibodies raised against the recombinant endobacterial CAT. CAT was detected in the endobacteria in the hypodermis of adult male and female
O.
volvulus, O. ochengi, O. gibsoni and
O.
fasciata. The endobacterial enzyme was also detected in onchocercal oocytes and all embryonic stages including intrauterine microfilariae as well as skin microfilariae.
O.
volvulus thus harbours
Wolbachia-like endosymbionts which are transovarially transmitted and show particular affinity for the hypodermal tissues of the lateral chords.