Phenanthrene degradation by bacteria of the genera <Emphasis Type="Italic">Pseudomonas</Emphasis> and <Emphasis Type="Italic">Burkholderia</Emphasis> in model soil systems

Degradation of phenanthrene by strains Pseudomona, Moscow, KMK, 2004simova, I.A. and Chernov, I.s putida BS3701 (pBS1141, pBS1142), Pseudomonas putida BS3745 (pBS216), and Burkholderia sp. BS3702 (pBS1143) were studied in model soil systems. The differences in accumulation and uptake rate of phenanthrene intermediates between the strains under study have been shown. Accumulation of 1-hydroxy-2-naphthoic acid in soil in the course of phenanthrene degradation by strain BS3702 (pBS1143) in a model system has been revealed. The efficiency of phenanthrene biodegradation was assessed using the mathematical model proposed previously for assessment of naphthalene degradation efficiency. The efficiency of degradation of both phenanthrene and the intermediate products of its degradation in phenanthrene-contaminated soil is expected to increase with the joint use of strains P. Putida BS3701 (pBS1141, pBS1142) and Burkholderia sp. BS3702 (pBS1143).     

Dioxygenases of chlorobiphenyl-degrading species <Emphasis Type="Italic">Rhodococcus wratislaviensis</Emphasis> G10 and chlorophenol-degrading species <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> 1CP induced in benzoate-grown cells and genes potentially involved in these processes

Dioxygenases induced during benzoate degradation by the actinobacterium Rhodococcus wratislaviensis G10 strain degrading haloaromatic compounds were studied. Rhodococcus wratislaviensis G10 completely degraded 2 g/liter benzoate during 30 h and 10 g/liter during 200 h. Washed cells grown on benzoate retained respiration activity for more than 90 days, and a high activity of benzoate dioxygenase was recorded for 10 days. Compared to the enzyme activities with benzoate, the activity of benzoate dioxygenases was 10-30% with 13 of 35 substituted benzoate analogs. Two dioxygenases capable of cleaving the aromatic ring were isolated and characterized: protocatechuate 3,4-dioxygenase and catechol 1,2-dioxygenase. Catechol inhibited the activity of protocatechuate 3,4-dioxygenase. Protocatechuate did not affect the activity of catechol 1,2-dioxygenase. A high degree of identity was shown by MALDI-TOF mass spectrometry for protein peaks of the R. wratislaviensis G10 and Rhodococcus opacus 1CP cells grown on benzoate or LB. DNA from the R. wratislaviensis G10 strain was specifically amplified using specific primers to variable regions of genes coding αand β-subunits of protocatechuate 3,4-dioxygenase and to two genes of theR. opacus 1CP coding catechol 1,2-dioxygenase. The products were 99% identical with the corresponding regions of the R. opacus 1CP genes. This high identity (99%) between the genes coding degradation of aromatic compounds in the R. wratislaviensis G10 and R. opacus 1CP strains isolated from sites of remote location (1400 km) and at different time (20-year difference) indicates a common origin of biodegradation genes of these strains and a wide distribution of these genes among rhodococci.     

Boosting fatty acid synthesis in <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> PD630 by overexpression of autologous thioesterases

Objectives

To explore the role of thioesterases in Rhodococcus opacus PD630 by endogenously overexpression in this bacteria for increased lipid production.

Results

Overexpression of four thioesterases from R. opacus PD630 in E. coli led to a 2- to 8-fold increase in C16:1 and C18:1 fatty acids while, when overexpressed in R. opacus PD630, only two recombinants had significant effect on the quantities and compositions of total fatty acid. The contents of total fatty acids (FAs) in two recombinants, pJTE2 (OPAG_00508 thioesterase) and pJTE4 (WP_012687673.1 thioesterase), were 400–460 mg/g (CDW) which is 1.5 times of wild-type strain PD630 (300-350 mg/g CDW), and 20–30 % (w/w) more than that of the control strain PDpJAM2 (330-370 mg/g CDW). The contents of 17:1 and 18:1 fatty acids increased by about 27 and 35 %, respectively, in pJTE2 and by 35 and 20 %, respectively, in pJTE4 compared with the control strain.

Conclusions

The engineered strains showed improved production of lipid (as total fatty acids), and could also tailor the composition of the fatty acid profile when cultured in mineral salts medium using glucose as sole carbon source.

     

Phenanthrene biodegradation and the interaction of <Emphasis Type="Italic">Pseudomonas putida</Emphasis> BS3701 and <Emphasis Type="Italic">Burkholderia</Emphasis> sp. BS3702 in plant rhizosphere

The interaction between the strains degrading polycyclic aromatic hydrocarbons, Pseudomonas putida BS3701 and Burkholderia sp. BS3702, was studied in the course of phenanthrene degradation in plant rhizosphere. Strain BS3702 was shown to accumulate 1-hydroxy-2-naphthoic acid (which is toxic for plants); it was then utilized by strain BS3701, which thereby increased the resistance of plants to the pollutant and to the toxic intermediate. With this type of interaction (cooperation), the efficiency of phenanthrene degradation was noted to decrease.     

Lipid metabolism of phenol-tolerant <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> strains for lignin bioconversion

Background

Lignin is a recalcitrant aromatic polymer that is a potential feedstock for renewable fuel and chemical production. Rhodococcus opacus PD630 is a promising strain for the biological upgrading of lignin due to its ability to tolerate and utilize lignin-derived aromatic compounds. To enhance its aromatic tolerance, we recently applied adaptive evolution using phenol as a sole carbon source and characterized a phenol-adapted R. opacus strain (evol40) and the wild-type (WT) strain by whole genome and RNA sequencing. While this effort increased our understanding of the aromatic tolerance, the tolerance mechanisms were not completely elucidated.

Results

We hypothesize that the composition of lipids plays an important role in phenol tolerance. To test this hypothesis, we applied high-resolution mass spectrometry analysis to lipid samples obtained from the WT and evol40 strains grown in 1 g/L glucose (glucose), 0.75 g/L phenol (low phenol), or 1.5 g/L phenol (high phenol, evol40 only) as a sole carbon source. This analysis identified?>?100 lipid species of mycolic acids, phosphatidylethanolamines (PEs), phosphatidylinositols (PIs), and triacylglycerols. In both strains, mycolic acids had fewer double bond numbers in phenol conditions than the glucose condition, and evol40 had significantly shorter mycolic acid chain lengths than the WT strain in phenol conditions. These results indicate that phenol adaptation affected mycolic acid membrane composition. In addition, the percentage of unsaturated phospholipids decreased for both strains in phenol conditions compared to the glucose condition. Moreover, the PI content increased for both strains in the low phenol condition compared to the glucose condition, and the PI content increased further for evol40 in the high phenol condition relative to the low phenol condition.

Conclusions

This work represents the first comprehensive lipidomic study on the membrane of R. opacus grown using phenol as a sole carbon source. Our results suggest that the alteration of the mycolic acid and phospholipid membrane composition may be a strategy of R. opacus for phenol tolerance.

     

Identification of inducible proteins in the phenanthrene degrader <Emphasis Type="Italic">Sphingobium chungbukense</Emphasis> DJ77 by 2-dimentional electrophoresis and liquid chromatography/tandem mass spectrometry

Sphingomonas chungbukensis DJ77 is a novel aromatic hydrocarbon-degrading bacterium capable of growing on phenanthrene as its sole source of carbon and energy. In this study, the protein expression profiles of S. chungbukensis DJ77 grown in the presence of phenanthrene were investigated by using two-dimensional gel electrophoresis (2-DE). Among 1000 protein spots visualized by 2-DE, the four proteins (i.e. 4-oxalocrotonate decarboxylase, 2-hydroxy-6-oxo-phenylhexa-2,4-dienoate hydrolase, glutathione S-transferase, and 2,3-dihydroxybiphenyl 1,2-dioxygenase) showing the significant upregulation by phenanthrene were identified by reversed-phase liquid chromatography–tandem mass spectrometry. Evidently, these proteins were involved in the metabolism of aromatic hydrocarbons. This can explain why S. chungbukensis DJ77 shows a significantly higher rate of phenanthrene consumption during the degradation process. The present analysis of proteomic responses and the detailed analysis results will be quite helpful to better understand the global physiology of S. chungbukensis DJ77, as proteome databases for various aromatic hydrocarbon-degrading strains have already been established.     

Production of single cell protein from agro-waste using <Emphasis Type="Italic">Rhodococcus opacus</Emphasis>

Livestock and fish farming are rapidly growing industries facing the simultaneous pressure of increasing production demands and limited protein required to produce feed. Bacteria that can convert low-value non-food waste streams into singe cell protein (SCP) present an intriguing route for rapid protein production. The oleaginous bacterium Rhodococcus opacus serves as a model organism for understanding microbial lipid production. SCP production has not been explored using an organism from this genus. In the present research, R. opacus strains DSM 1069 and PD630 were fed three agro-waste streams: (1) orange pulp, juice, and peel; (2) lemon pulp, juice, and peel; and (3) corn stover effluent, to determine if these low-cost substrates would be suitable for producing a value-added product, SCP for aquafarming or livestock feed. Both strains used agro-waste carbon sources as a growth substrate to produce protein-rich cell biomass suggesting that that R. opacus can be used to produce SCP using agro-wastes as low-cost substrates.     

Phenol degradation by <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> strain 1G

During cultivation in a liquid medium, the bacterium Rhodococcus opacus 1G was capable of growing on phenol at a concentration of up to 0.75 g/l. Immobilization of Rhodococcus opacus 1G had a positive effect on cell growth in the presence of phenol at high concentrations. The substrate at concentrations of 1.0 and 1.5 g/l was completely utilized over 24 and 48 h, respectively. The key enzymes of phenol degradation (two catechol 1,2-dioxygenases and muconate cycloisomerase) were isolated. One of the dioxygenases was very unstable. By substrate specificity, another enzyme belonged to catechol 1,2-dioxygenases of the classical ortho-pathway. Chlorocatechols and chlorophenols served as competitive inhibitors of catechol 1,2-dioxygenases. The inhibitory effect of other aromatic compounds was less significant. Our results suggest that this strain holds promise for bioremediation of phenol wastewater.     

Simultaneous Biodegradation of Polyaromatic Hydrocarbons by a <Emphasis Type="Italic">Stenotrophomonas</Emphasis> sp: Characterization of <Emphasis Type="Italic">nid</Emphasis> Genes and Effect of Surfactants on Degradation

A polyaromatic hydrocarbon degrading bacterium was isolated from a petroleum contaminated site and designated as Stenotrophomonas sp. strain IITR87. It was found to utilize pyrene, phenanthrene and benzo(a)pyrene as sole carbon source, but not anthracene, chrysene and fluoranthene. Gas chromatography and mass spectroscopy analysis resulted in identification of pyrene metabolites namely monohydroxypyrene, 4-oxa-pyrene-5-one, dimethoxypyrene and monohydroxyphenanthrene. Southern hybridization using naphthalene dioxygenase gene (nidA) as probe against the DNA of strain IITR87 revealed the presence of nidA gene. PCR analysis suggests dispersed occurrence of nid genes in the genome instead of a cluster as reported in a PAH-degrading Mycobacterium vanbaalenii PYR-1. The nid genes in strain IITR87, dioxygenase large subunit (nidA), naphthalene dioxygenase small subunit (nidB) and aldehyde dehydrogenase gene (nidD) showed more than 97 % identity to the reported nid genes from Mycobacterium vanbaalenii PYR-1. Most significantly, the biodegradation of PAHs was enhanced 25–60 % in the presence of surfactants rhamnolipid and Triton X-100 due to increased solubilization and bioavailability. These results could be useful for the improved biodegradation of high-molecular-weight PAHs in contaminated habitats.     

Epiphytic microorganisms degrading aromatic hydrocarbons from the phyllosphere of urban woody plants

From the leaves of three urban trees (Tilia sp., Acer sp., and Fraxinus sp.), 180 strains degrading phenanthrene, naphthalene, and salicylate were isolated by direct plating and enrichment cultures. The leaves of each tree species were characterized by a specific profile of aromatic hydrocarbon-degrading microflora. Members of the type Actinobacteria were predominant in the case of direct plating on media with phenanthrene and naphthalene. Enrichment cultures with phenanthrene and salicylate were shown to yield microbial consortia, the composition of which changed with time. Members of the type Proteobacteria were predominant in these consortia. No plasmids of polycyclic aromatic hydrocarbon degradation of the P-7 and P-9 incompatibility groups were revealed in the studied strains.     

A consortium of immobilized rhodococci for oilfield wastewater treatment in a column bioreactor

The possible use of a consortium of actinobacteria from the genus Rhodococcus immobilized on a polymeric carrier has been investigated for oilfield wastewater treatment in a bioreactor. It has been found that Rhodococcus opacus IEGM 263 and Rhodococcus ruber IEGM 231 cells remain viable at high concentrations of mineral salts in water and are able to oxidize oil hydrocarbons up to 62–81%. It has been shown that the consortium of rhodococci was more efficient in the elimination of hydrocarbons from wastewater than monocultures.     

Diversity and symbiotic effectiveness of <Emphasis Type="Italic">Adesmia</Emphasis> spp. root nodule bacteria in central and southern Chile

Legumes in the genus Adesmia are wild species with forage and medicinal potential. Their nitrogen fixation efficiency depends on their association with soil bacteria known as rhizobia. The aim of this work was to assess the diversity and symbiotic effectiveness of root nodule bacteria from Adesmia boronioides, Adesmia emarginata and Adesmia tenella from different regions of Chile. Adesmia spp. nodules were collected from seven sites obtaining 47 isolates, which resulted in 19 distinct strains. The diversity of the strains was determined via partial sequencing of the dnaK, 16srRNA and nodA genes. The strains were authenticated as root nodule bacteria on their original host and assessed for symbiotic effectiveness on A. emarginata and A. tenella. The strains from Adesmia tenella clustered within the Mesorhizobium clade. Adesmia boronioides nodulated with Mesorhizobium sp., Rhizobium leguminosarum and Bradyrhizobium sp. The rhizobia from A. emarginata were identified as Burkholderia spp, which was symbiotically ineffective on this species and on A. tenella. Strains isolated from Adesmia emarginata nodules, but unable to induce nodulation, were identified as Labrys methylaminiphilus. Labrys strain AG-49 significantly increased root dry weight in A. emarginata. The nodA genes from Adesmia strains were unique and correlated to legume host. A. emarginata was effectively nodulated by Bradyrhizobium AG-64 and A. tenella by Mesorhizobium strains AG-51 and AG.52. It is concluded that Adesmia emarginata, A. tenella and A. boronioides are associated to diverse bacterial symbionts and selection of an effective inoculant is a key step to assist Adesmia spp. adaptation and restoration.     

Cyclohexanone-induced stress metabolism of <Emphasis Type="Italic">Escherichia coli</Emphasis> and <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis>

Solvent stress occurs during whole-cell biocatalysis of organic chemicals. Organic substrates and/or products may accumulate in the cellular membranes of whole cells, causing structural destabilization of the membranes, which leads to disturbances in cellular carbon and energy metabolism. Here, we investigate the effect of cyclohexanone on carbon metabolism in Escherichia coli BL21 and Corynebacterium glutamicum ATCC13032. Adding cyclohexanone to the culture medium (i.e., glucose mineral medium) resulted in a decreased specific growth rate and increased cellular maintenance energy in both strains of bacteria. Notably, carbon metabolism, which is mainly involved to increase cellular maintenance energy, was very different between the bacteria. Carbon flux into the acetic acid fermentation pathway was dominantly enhanced in E. coli, whereas the TCA cycle appeared to be activated in C. glutamicum. In fact, carbon flux into the TCA cycle in E. coli appeared to be reduced with increasing amounts of cyclohexanone in the culture medium. Metabolic engineering of E. coli cells to maintain or improve TCA cycle activity and, presumably, that of the electron transport chain, which are involved in regeneration of cofactors (e.g., NAD(P)H and ATP) and formation of toxic metabolites (e.g., acetic acid), may be useful in increasing solvent tolerance and biotransformation of organic chemicals (e.g., cyclohexanone).     

Dearomatization of diesel oil using <Emphasis Type="Italic">Pseudomonas</Emphasis> sp.

Objectives

To improve the quality of diesel fuel via removal of aromatic compounds using Pseudomonas sp.

Results

In the present study Pseudomonas sp. was able to remove 94% of fluorene, 59% of phenanthrene, 49% of anthracene, 52% of fluoranthene, 45% of pyrene and 75% carbazole present in diesel oil. Additionally, it also does not affect the aliphatic content of fuel thus maintaining the carbon backbone of the fuel.

Conclusions

Pseudomonas sp. is a potential biocatalyst that can be used in the refining industry.

     

<Emphasis Type="Italic">glyA</Emphasis> gene knock-out in <Emphasis Type="Italic">Escherichia coli</Emphasis> enhances L-serine production without glycine addition

In E. coli, glyA encodes for serine hydroxymethyltransferase (SHMT), which converts L-serine to glycine. When engineering L-serine-producing strains, it is therefore favorable to inactivate glyA to prevent L-serine degradation. However, most glyA knockout strains exhibit slow cell growth because of the resulting lack of glycine and C₁ units. To overcome this problem, we overexpressed the gcvTHP genes of the glycine cleavage system (GCV), to increase the C₁ supply before glyA was knocked out. Subsequently, the kbl and tdh genes were overexpressed to provide additional glycine via the L-threonine degradation pathway, thus restoring normal cell growth independent of glycine addition. Finally, the plasmid pPK10 was introduced to overexpress pgk, serA ^Δ197 , serC and serB, and the resulting strain E4G2 (pPK10) accumulated 266.3 mg/L of L-serine in a semi-defined medium without adding glycine, which was 3.18-fold higher than the production achieved by the control strain E3 (pPK10). This strategy can accordingly be applied to disrupt the L-serine degradation pathway in industrial production strains without causing negative side-effects, ultimately making L-serine production more efficient.     

Engineering of an L-arabinose metabolic pathway in <Emphasis Type="Italic">Rhodococcus jostii</Emphasis> RHA1 for biofuel production

The oleaginous bacterium, Rhodococcus jostii RHA1 has attracted considerable attention due to its capability to accumulate significant levels of triacylglycerol as renewable hydrocarbon. To enable the strain to utilize arabinose derived from lignocellulosic biomass, the metabolic pathway of L-arabinose utilization was introduced into R. jostii RHA1 by heterogenous expression of the operon, araBAD from Escherichia coli. The results showed that recombinant bearing araBAD could grow on L-arabinose as the sole carbon source, and additional expression of araFGH encoding the arabinose transporter from E. coli could improve the cell biomass yield from high contents of arabinose. We further increased the content of lipid produced from arabinose in the recombinants from 47.9 to 56.8 % of the cell dry weight (CDW) by overexpression of a gene, atf1 encoding a diglyceride acyltransferase from R. opacus PD630. This work demonstrated the feasibility of producing lipid from arabinose by genetic modification of the rhodococci strain.     

Secretion of Proteinases with Fibrinolytic Activity by Micromycetes of the Genus <Emphasis Type="Italic">Aspergillus</Emphasis>

Proteolytic activity of extracellular enzymes of 11 strains of different Aspergillus species was studied. Comparison of the enzymatic indices of strains grown on agar medium containing either casein or fibrin allowed the selection of the strain Aspergillus terreus 2 as a promising producer of fibrinolytic proteases. It was found that A. terreus 2 proteinases demonstrated maximum activity at pH 8.0. The highest values of fibrinolytic and total proteolytic activities expressed in U_Tyr (amount of micromoles of tyrosine released from fibrin or casein for 1 min) were 34.0 and 358.3, respectively. Maximum activities were detected when growing the producer on a medium containing only amine nitrogen sources (fish flour hydrolysate and peptone); however, the amount of extracellular protein and the specific fibrinolytic and total proteolytic activities were greater in the medium containing both mineral and amine nitrogen sources (fish flour hydrolysate and sodium nitrate) than in the medium containing only fish flour hydrolysate and peptone as nitrogen sources.     

Overexpression of a phosphatidic acid phosphatase type 2 leads to an increase in triacylglycerol production in oleaginous <Emphasis Type="Italic">Rhodococcus</Emphasis> strains

Oleaginous Rhodococcus strains are able to accumulate large amounts of triacylglycerol (TAG). Phosphatidic acid phosphatase (PAP) enzyme catalyzes the dephosphorylation of phosphatidic acid (PA) to yield diacylglycerol (DAG), a key precursor for TAG biosynthesis. Studies to establish its role in lipid metabolism have been mainly focused in eukaryotes but not in bacteria. In this work, we identified and characterized a putative PAP type 2 (PAP2) encoded by the ro00075 gene in Rhodococcus jostii RHA1. Heterologous expression of ro00075 in Escherichia coli resulted in a fourfold increase in PAP activity and twofold in DAG content. The conditional deletion of ro00075 in RHA1 led to a decrease in the content of DAG and TAG, whereas its overexpression in both RHA1 and Rhodococcus opacus PD630 promoted an increase up to 10 to 15 % by cellular dry weight in TAG content. On the other hand, expression of ro00075 in the non-oleaginous strain Rhodococcus fascians F7 promoted an increase in total fatty acid content up to 7 % at the expense of free fatty acid (FFA), DAG, and TAG fractions. Moreover, co-expression of ro00075/atf2 genes resulted in a fourfold increase in total fatty acid content by a further increase of the FFA and TAG fractions. The results of this study suggest that ro00075 encodes for a PAP2 enzyme actively involved in TAG biosynthesis. Overexpression of this gene, as single one or with an atf gene, provides an alternative approach to increase the biosynthesis and accumulation of bacterial oils as a potential source of raw material for biofuel production.     

The <Emphasis Type="Italic">Adh</Emphasis> locus and adaptation of <Emphasis Type="Italic">cn</Emphasis> and <Emphasis Type="Italic">vg</Emphasis> mutants in experimental populations of <Emphasis Type="Italic">Drosophila melanogaster MEIG</Emphasis>

A complex study on the adaptation of cn and vn mutants and the allozymes of alcoholdehydrogenase (ADH) was carried out in initially pure lines, and their panmixia populations during exchange of the mutant genotype with that of wild-type flies (C-S) and D) through saturating crossings. The relative adaptation of the genotypes was estimated by their effect on reproductive efficiency in the experimentally obtained population. Fecundity, lifespan, and the resistance of the studied genotypes to hyperthermia were investigated individually. It was shown that the high level of adaptation of the cn mutants and the low level of adaptation of the vg mutants was correlated with the presence of different ADH allozymes. In the studied population, the F-allozyme of ADH accompanied the vg mutation, while the S-allozyme of the enzyme was detected in cn mutants. Saturating crossings of C-S(Adh ^S)×vg(Adh ^F) and D(Adh ^F) × cn(Adh ^S), along with the parallel determination of the allele composition of the Adh locus, demonstrated that the complete substitution of the F-allozyme of ADH in the vg mutants by the S-allozyme in D flies, as well as the substitution of the S-allozyme of ADH in the cn mutants by the F-allozyme in D flies was realized only after the 15th–20th backcrosses. These results favor the coadaptation of cn and vg marker genes with alleles of the Adh locus and indicate the important role of the latter in the adaptation of genotypes. In the studied population, selection acted primarily against the vg mutants, which were inferior to the cn mutants, and heterozygote genotypes in indices of the main adaptation components.     

The regulation of peroxisomal matrix enzymes (alcohol oxidase and catalase) formation by the product of the gene <Emphasis Type="Italic">Mth1</Emphasis> in methylotrophic yeast <Emphasis Type="Italic">Pichia methanolica</Emphasis>

Two independent mutant strains of methylotrophic yeast Pichia methanolica (mth1 arg1 and mth2 arg4) from the initial line 616 (ade1 ade5) were investigated. The mutant strains possessed defects in genes MTH1 and MTH2 which resulted in the inability to assimilate methanol as a sole carbon source and the increased activity of alcohol oxidase (AO). The function of the AUG2 gene encoding one of the subunits of AO and CTA1, a probable homolog of peroxisomal catalase of Saccharomyces cereviseae, was investigated by analyses of the molecular forms of isoenzymes. It was shown that optimal conditions for the expression of the AUG2 gene on a medium supplemented with 3% of methanol leads to an increasing synthesis of peroxisomal catalase. The mutant mth1 possessed a dominant formation of AO isoform with electrophoretic mobility which is typical for isogenic form 9, the product of the AUG2 gene, and a decreased level of peroxisomal catalase. The restoration of growth of four spontaneous revertants of the mutant mth1 (Rmth1) on the methanol containing medium was accompanied by an increase in activity of AO isogenic form 9 and peroxisomal catalase. The obtained results confirmed the functional continuity of the structural gene AUG2 in mutant mth1. The correlation of activity of peroxisomal catalase and AO isogenic form 1 in different conditions evidenced the existence of common regulatory elements for genes AUG2 and CTA1 in methilotrophic yeast Pichia methanolica.