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Fungal Carbonyl Sulfide Hydrolase of Trichoderma harzianum Strain THIF08 and Its Relationship with Clade D β-Carbonic Anhydrases.

ABSTRACT: Carbonyl sulfide (COS) is the most abundant and long-lived sulfur-containing gas in the atmosphere. Soil is the main sink of COS in the atmosphere and uptake is dominated by soil microorganisms; however, biochemical research has not yet been conducted on fungal COS degradation. COS hydrolase (COSase) was purified from Trichoderma harzianum strain THIF08, which degrades COS at concentrations higher than 10,000 parts per million by volume from atmospheric concentrations, and its gene cos (492 bp) was cloned. The recombinant protein purified from Escherichia coli expressing the cos gene converted COS to H₂S. The deduced amino acid sequence of COSase (163 amino acids) was assigned to clade D in the phylogenetic tree of the β-carbonic anhydrase (β-CA) family, to which prokaryotic COSase and its structurally related enzymes belong. However, the COSase of strain THIF08 differed from the previously known prokaryotic COSase and its related enzymes due to its low reactivity to CO₂ and inability to hydrolyze CS₂. Sequence comparisons of the active site amino acids of clade D β-CA family enzymes suggested that various Ascomycota, particularly Sordariomycetes and Eurotiomycetes, possess similar enzymes to the COSase of strain THIF08 with >80% identity. These fungal COSase were phylogenetically distant to prokaryotic clade D β-CA family enzymes. These results suggest that various ascomycetes containing COSase contribute to the uptake of COS by soil.

SUBMITTER: Masaki Y

PROVIDER: S-EPMC8209446 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Fungi have the capacity to assimilate a diverse range of both inorganic and organic sulfur compounds. It has been recognized that all sulfur sources taken up by fungi are in soluble forms. In this study, we present evidence that fungi can utilize gaseous carbonyl sulfide (COS) for the assimilation of a sulfur compound. We found that the filamentous fungus Trichoderma harzianum strain THIF08, which has constitutively high COS-degrading activity, was able to grow with COS as the sole sulfur source. Cultivation with 34S-labeled COS revealed that sulfur atom from COS was incorporated into intracellular metabolites such as glutathione and ergothioneine. COS degradation by strain THIF08, in which as much of the moisture derived from the agar medium as possible was removed, indicated that gaseous COS was taken up directly into the cell. Escherichia coli transformed with a COS hydrolase (COSase) gene, which is clade D of the β-class carbonic anhydrase subfamily enzyme with high specificity for COS but low activity for CO2 hydration, showed that the COSase is involved in COS assimilation. Comparison of sulfur metabolites of strain THIF08 revealed a higher relative abundance of reduced sulfur compounds under the COS-supplemented condition than the sulfate-supplemented condition, suggesting that sulfur assimilation is more energetically efficient with COS than with sulfate because there is no redox change of sulfur. Phylogenetic analysis of the genes encoding COSase, which are distributed in a wide range of fungal taxa, suggests that the common ancestor of Ascomycota, Basidiomycota, and Mucoromycota acquired COSase at about 790-670 Ma.IMPORTANCEThe biological assimilation of gaseous CO2 and N2 involves essential processes known as carbon fixation and nitrogen fixation, respectively. In this study, we found that the fungus Trichoderma harzianum strain THIF08 can grow with gaseous carbonyl sulfide (COS), the most abundant and ubiquitous gaseous sulfur compound, as a sulfur source. When the fungus grew in these conditions, COS was assimilated into sulfur metabolites, and the key enzyme of this assimilation process is COS hydrolase (COSase), which specifically degrades COS. Moreover, the pathway was more energy efficient than the typical sulfate assimilation pathway. COSase genes are widely distributed in Ascomycota, Basidiomycota, and Mucoromycota and also occur in some Chytridiomycota, indicating that COS assimilation is widespread in fungi. Phylogenetic analysis of these genes revealed that the acquisition of COSase in filamentous fungi was estimated to have occurred at about 790-670 Ma, around the time that filamentous fungi transitioned to a terrestrial environment.

Project description:In the mosquito midgut, luminal pH regulation and cellular ion transport processes are important for the digestion of food and maintenance of cellular homeostasis. pH regulation in the mosquito gut is affected by the vectorial movement of the principal ions including bicarbonate/carbonate and protons. As in all metazoans, mosquitoes employ the product of aerobic metabolism carbon dioxide in its bicarbonate/carbonate form as one of the major buffers of cellular and extracellular pH. The conversion of metabolic carbon dioxide to bicarbonate/carbonate is accomplished by a family of enzymes encoded by the carbonic anhydrase gene family. This study characterizes Aedes aegypti carbonic anhydrases using bioinformatic, molecular, and immunohistochemical methods. Our analyses show that there are fourteen Aedes aegypti carbonic anhydrase genes, two of which are expressed as splice variants. The carbonic anhydrases were classified as either integral membrane, peripheral membrane, mitochondrial, secreted, or soluble cytoplasmic proteins. Using polymerase chain reaction and Western blotting, one of the carbonic anhydrases, Aedes aegypti carbonic anhydrase 9, was analyzed and found in each life stage, male/female pupae, male/female adults, and in the female posterior midgut. Next, carbonic anhydrase 9 was analyzed in larvae and adults using confocal microscopy and was detected in the midgut regions. According to our analyses, carbonic anhydrase 9 is a soluble cytoplasmic enzyme found in the alimentary canal of larvae and adults and is expressed throughout the life cycle of the mosquito. Based on previous physiological analyses of adults and larvae, it appears AeCA9 is one of the major carbonic anhydrases involved in producing bicarbonate/carbonate which is involved in pH regulation and ion transport processes in the alimentary canal. Detailed understanding of the molecular bases of ion homeostasis in mosquitoes will provide targets for novel mosquito control strategies into the new millennium.

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Fungal Carbonyl Sulfide Hydrolase of Trichoderma harzianum Strain THIF08 and Its Relationship with Clade D β-Carbonic Anhydrases.

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