Effect of chemical water pollutants on the growth of two Paramecium species (Paramecium caudatum and Paramecium trichium) were examined. The chemicals used as model chemical pollutants include organic solvents, potential carcinogens, mutagens, metabolic modulators, herbicides, insecticides, fungicides, antimicrobials, heavy metals, and heavy metal-containing chemicals. In this study, the IC 50 values indicating the concentration of substances inhibiting the proliferation of Paramecium cells by 50% were used instead of the LD 50 value, which indicates the dose of substances killing half the population of organisms, since the former is a more sensitive parameter for assessing the toxicity of substances at lower concentrations. Among 25 chemicals examined, di-(2-ethylhexyl)phthalate, potassium dichromate, 2,4-dichlorophenoxy acetic acid, and paraquat stimulated the growth of paramecia depending on the concentrations used. Dimethyl sulfoxide and formaldehyde were shown to be inert to paramecia in the range of concentrations (up to 1%, v/v) used here. Other chemicals were shown to inhibit the growth of the paramecia and thus the IC 50 values for those chemicals were determined. Our data presented here may be a useful reference for assessing the impact of water pollutants on aqueous microecosystems consisting of various microorganisms including protozoa.
A protease in the culture medium of Tetrahymena pyriformis was purified to homogeneity. The purified protease had an apparent molecular mass of 28 kDa on SDS/PAGE. The amino acid sequences of the N-terminal and internal peptides of the protease showed complete identity with those of tetrain, an enzyme previously reported as a Tetrahymena cysteine protease but not characterized in detail. Two overlapping cDNA clones for tetrain were sequenced, and the nucleotide sequence predicts that these clones encode a 330-amino acid protein composed of a 16-residue N-terminal signal sequence followed by a 103-residue propeptide and a 211-residue mature protease. The primary structure and enzymatic properties support the conclusion that tetrain belongs to the cathepsin L subfamily. Immunoblotting analyses showed that mature tetrain was found exclusively in the culture medium. Immunofluorescence microscopy demonstrated that tetrain was concentrated in or around the food vacuoles of cells in the late logarithmic phase, but the staining of food vacuoles was not obvious in the stationary phase. These results suggest that tetrain is synthesized at the logarithmic phase and is secreted into the culture medium as a mature form.Keywords : cathepsin L; cysteine protease; growth phase ; Tetrahymena; tetrain.Proteases are expressed in various cells and tissues of nu-increased during cell growth [6]. The protease activity was partially purified, identified as a member of the lysosomal cysteine merous species of living organisms and are classified into large groups including the cysteine, serine, aspartic, and met-proteases by amino acid sequencing, and designated as tetrain, a Tetrahymena cysteine protease. Because tetrain is present in alloproteases. Cysteine proteases consist of two diverse groups of enzymes which localize mainly to digestive vacuoles or lyso-the culture medium at higher levels in the stationary phase than in the logarithmic phase, it becomes interesting to question the somes (cathepsins) [1] and the cytosol (calpain) [2]. These enzymes have a variety of biological functions such as intracellular biological significance of protease secretion for free-living ciliates. protein degradation or the precise processing of precursor proteins. In some cases, cysteine proteases are secreted into the Tetrahymena possesses at least two independent secretory pathways, one resulting in the secretion of mucocysts [7] and extracellular environment. It has been speculated that cathepsins secreted by certain tumor cells play an active extracellular role another resulting in the secretion of lysosomal enzymes [8]. Because of specialized cell-surface domains such as the cytoproct in cancer metastasis and tissue remodeling [3]. In some parasites, it has been suggested that secreted cysteine proteases par-or a well-developed lysosome-phagosome system [9], Tetrahymena has frequently been used as a material for studying these ticipate in the supply of nutritional requirements or in damaging host tissue [4,5]. Previously, we have found that pr...
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