Trichomonas vaginalis acidic phospholipase A2: isolation and partial amino acid sequence

Abstract

Sexually transmitted diseases are a major cause of acute disease worldwide, and trichomoniasis is the most common and curable disease, generating more than 170 million cases annually worldwide. Trichomonas vaginalis is the causal agent of trichomoniasis and has the ability to destroy in vitro cell monolayers of the vaginal mucosa, where the phospholipases A 2 (PLA 2 ) have been reported as potential virulence factors. These enzymes have been partially characterized from the subcellular fraction S30 of pathogenic T. vaginalis strains. The main objective of this study was to purify a phospholipase A 2 from T. vaginalis, make a partial characterization, obtain a partial amino acid sequence, and determine its enzymatic participation as hemolytic factor causing lysis of erythrocytes. Trichomonas S30, RF30 and UFF30 sub-fractions from GT-15 strain have the capacity to hydrolyze [2-14 C-PA]-PC at pH 6.0. Proteins from the UFF30 sub-fraction were separated by affinity chromatography into two eluted fractions with detectable PLA A 2 activity. The EDTA-eluted fraction was analyzed by HPLC using on-line HPLC-tandem mass spectrometry and two protein peaks were observed at 8.2 and 13 kDa. Peptide sequences were identified from the proteins present in the eluted EDTA UFF30 fraction; bioinformatic analysis using Protein Link Global Server charged with T. vaginalis protein database suggests that eluted peptides correspond a putative ubiquitin protein in the 8.2 kDa fraction and a phospholipase preserved in the 13 kDa fraction. The EDTA-eluted fraction hydrolyzed [2-14 C-PA]-PC lyses erythrocytes from Sprague-Dawley in a time and dose-dependent manner. The acidic hemolytic activity decreased by 84% with the addition of 100 µM of Rosenthal's inhibitor.

Phospholipases A and Lysophospholipases in protozoan parasites

“…A PLA2 was isolated from a subcellular fraction, but solid functional genomics proofs need to be added. This eluted fraction caused hemolysis and was inhibited by Rosenthal's inhibitor [ 106 ]. The genome of T. vaginalis was sequenced in 2007 [ 107 ], but no PLA gene has been identified since then.…”

Recent Advances in the Trichomonas vaginalis Field

“…It is important to note that cell adhesion is an absolutely necessary prerequisite for the lysis of host cells by Tv 38 . After cell adhesion has taken place, several factors are assumed to be involved in host cell lysis, including metalloproteases 39 , 40 , cysteine proteases 41 – 43 , a rhomboid protease (TvROM1) 44 , and phospholipase A2 45 . Tv also secretes a migration inhibition factor (TvMIF) 46 which can replace human migration factor (HuMIF) to trigger proinflammatory cytokine release.…”

“…Binding is accomplished by several surface proteins and other surface molecules that bind to a structure on the host’s cell surface. Among these are lipoglycan 27 , BspA 28 , 30 , tetraspanins 28 , 45 , 46 and several others, such as glyceraldehyde 3-phosphate dehydrogenase 32 , enolase 33 , and succinyl-CoA synthetase 34 on the Tv surface, galectins-1 and -3 28 on the host cell surface, and fibronectin 32 in the extracellular matrix. Several Tv factors necessary for adhesion to the host epithelium reach the Tv surface or the epithelium surface via exosomes 31 ( 2 ).…”

Development of a convenient detection method for Trichomonas vaginalis based on loop-mediated isothermal amplification targeting adhesion protein 65

“…According to statistics, more than 170 million people were infected with the disease every year in the world, and the infection rate of people around the world was different and had an increasing trend [9]. In the United States, nearly 5 million people were infected with T. vaginalis every year, while the infection rate was 24.3% in Japan, 23.8% in Uganda and 18.0% in South Africa [10].…”