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Microbiology and Biotechnology Letters

Genome Report(Note)

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Genome Report  |  Genome Report

Microbiol. Biotechnol. Lett. 2023; 51(4): 531-534

https://doi.org/10.48022/mbl.2310.10004

Received: October 5, 2023; Accepted: November 7, 2023

Draft Genome Sequence of the Yeast Strain Hormonema macrosporum POB-4, which Produces the Biosurfactant Glycocholic Acid

Parthiban Subramanian1, Jeong-Seon Kim2, Jun Heo2, and Yiseul Kim2*

1National Agrobiodiversity Center, National Institute of Agricultural Sciences, Rural Development Administration, Wanju 55365, Republic of Korea
2Agricultural Microbiology Division, National Institute of Agricultural Sciences, Rural Development Administration, Wanju 55365, Republic of Korea

Correspondence to :
Yiseul Kim,        dew@korea.kr

Abstract

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We report the draft genome sequence of the yeast strain Hormonema macrosporum POB-4, capable of producing the biosurfactant glycocholic acid, one of the bile acids. A majority of genes with known function were associated with metabolism and transport of amino acid and carbohydrate as well as secondary metabolites biosynthesis, transport, and catabolism. We observed genes of eleven C-N hydrolases and two CoA transferases which have been reported to be involved in the biosynthesis of glycocholic acid. Further experimental studies can help to elucidate the specific genes responsible for biosurfactant production in strain POB-4.

Keywords: Yeast, Hormonema macrosporum, biosurfactant, genome

Body

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With an aim of supporting the sustainable development, there has been a growing interest in microorganisms with industrial potential to reduce overuse of environment-polluting synthetic materials. We report here, the genomic information of biosurfactant, glycocholic acid producing yeast Hormonema macrosporum POB-4, isolated from flower pollen in the Republic of Korea. In order to study the metabolic potential of strain POB-4, it was subject to genome sequencing. The genome of strain POB-4 was sequenced using a combination of HiFi sequencing (Sequel II System) and Illumina HiSeq X-ten (Illumina, USA) platforms by Macrogen Inc., (Republic of Korea). Genome assembly application from PacBio was used to generate high quality de novo assemblies using HiFi reads. Briefly, PanCake 1.1.2 [1] was used to overlap the reads followed by Nighthawk to phase the overlapped reads. After the removal of chimeras and duplicates from the overlapped reads, a string graph was constructed generating primary contigs as well as haplotigs. The primary contigs and haplotigs were polished using Racon 1.5.0 [2] with phased reads. To remove haplotype duplications from the primary contig set, purge_dups was used for retrieving potential haplotype duplications and move them to the haplotig set. The generated genome assembly was subject to further analyses.

Unless otherwise specified, all further analyses were carried out on the Galaxy Web server (https://usegalaxy.org). Completeness assessment of genome assembly was examined using BUSCO 4.1.4 [3] with the lineage dataset dothideomycetes_odb10. Repetitive elements were studied using RepeatMasker 4.1.5 followed by gene prediction using AUGUSTUS software with Neurospora crassa as a model for training [4]. For quality assessment of gene prediction, annotations of strain POB-4 using the different tools, AUGUSTUS and Maker, were compared via genome annotation statistics tool available at the Galaxy Web server (Table 1). Subsequently, the functional annotation of the data from AUGUSTUS and MAKER was carried out using eggNOG-mapper 2 [5]. During mapping, the query genome was screened for Clusters of Orthologous Genes (COGs), Gene Ontology (GO terms), Carbohydrate-Active enZYmes (CAZy), and Pfam. The genome was also queried at the main pathway databases, including KEGG and PANTHER using KOBAS 2.0 to study functional metabolism of genes [6]. Secondary metabolite production by strain POB-4 was analyzed using fungal version of antiSMASH 7.0 [7] (https://fungismash.secondarymetabolites.org/#!/start). The raw sequencing data of strain POB-4 was deposited in GenBank with the accession number of SRX21170146 and SRX21170147.

Table 1 . Statistics of gene prediction using the different programs.

ATTRIBUTEAUGUSTUSMAKER
Contigs1616
Number of genes predicted8,6409,854
Number of transcripts predicted8,6409,854
Complete BUSCOs3,3523,261
Missing BUSCOs291130
Number of selected queries by7,1338,455
EggNOG-mapper(82.6%)(85.8%)
Single EggNOG6,7586,791
Multi EggNOG375929
Pfam hits*6,4617,613
GO hits*3,5404,021
EC hits*1,7532,012
CAZy hits*137162

*Number of predicted genes that contain at least one Pfam domain, one GO term, one enzyme, and one CAZy hit.

The genome assembly was 28.4 Mb (28,419,067 bp) in size and consisted of 16 scaffolds with N50 value of 2.2Mb (2,213,373 bp). The largest scaffold was 2,872,214 bp long and the shortest was 53,043 bp long. The GC content was estimated to be 49.8%. Completeness of the genome assembly was 97.3%, showing the following profile C: 97.3% [S: 96.9%, D: 0.4%], F: 0.2%, M: 2.5%, n: 3786 when dothideomycetes_odb10 dataset was used as reference. Analysis of repetitive elements exhibited very few repeat elements in the genome (0.92%). Functional annotation using outputs from AUGUSTUS (8,640 predicted genes) and MAKER (9,854 predicted genes) resulted in identification of functional traits of the coding sequences (Table 2). Strain POB-4 contained a majority of genes for metabolism and transport of amino acid as well as carbohydrate followed by secondary metabolite biosynthesis, transport, and catabolism. Analysis using fungal antiSMASH resulted in three hits, namely melanin (100% match, contig 6), neosatorin (52% match, contig 1), and polyketide synthesis (33% match, contig 10).

Table 2 . Metabolism related genes of the yeast strain Hormonema macrosporum POB-4.

COG CATAGORIESAUGUSTUSMAKER
INFORMATION STORAGE AND PROCESSING1,0301,208
[J] Translation, ribosomal structure and biogenesis319355
[A] RNA processing and modification272302
[K] Transcription206275
[L] Replication, recombination and repair155184
[B] Chromatin structure and dynamics7892
CELLULAR PROCESSES AND SIGNALING1,282988
[D] Cell cycle control, cell division, chromosome partitioning8290
[Y] Nuclear structure55
[V] Defense mechanisms4759
[T] Signal transduction mechanisms234277
[M] Cell wall/membrane/envelope biogenesis4754
[N] Cell motility33
[Z] Cytoskeleton7585
[W] Extracellular structures45
[U] Intracellular trafficking, secretion, and vesicular transport353410
[O] Posttranslational modification, protein turnover, chaperones4320
METABOLISM2,1682,491
[C] Energy production and conversion283314
[G] Carbohydrate transport and metabolism432511
[E] Amino acid transport and metabolism398458
[F] Nucleotide transport and metabolism7888
[H] Coenzyme transport and metabolism141165
[I] Lipid transport and metabolism242280
[P] Inorganic ion transport and metabolism205229
[Q] Secondary metabolites biosynthesis, transport and catabolism389446
POORLY CHARACTERIZED1,7012,104
[R] General function prediction only00
[S] Function unknown1,7012,104
No hits577735


The compound glycocholic acid, identified as the major component of the biosurfactant by strain POB-4 during HPLC and NMR analyses, is one of the bile acids (patent application number 10-2023-0147900). Microbial production of bile acid conjugates such as glycocholic acid has been documented previously. Two studies have shown the production of glycocholic acid by fungus Penicillum sp., which belongs to subdivision Pezizomycotina of Ascomycota same as strain POB-4 [8, 9]. Moreover, bacterial strains with marine origin as well as gut bacteria have been reported to produce bile acid conjugates [10−12]. In particular, Garcia et al. proposed amino acid N-acyltransferases as a mechanism for the production of microbially conjugated bile acids such as glycocholic acid [12]. In the KEGG database, glycocholic acid (compound C01921) leads to 1,027 reported genes, which are orthologues of two genes, namely bile acid-CoA: amino acid Nacyltransferase (K00659) and choloylglycine hydrolase (EC 3.5.1.24) or linear amide C-N hydrolase (K01442). Although genes associated with neither bile acid-CoA: amino acid N-acyltransferases nor choloylglycine hydrolase were detected, we identified eleven genes for C-N hydrolase as well as two genes for acyl-CoA thioesterase (EC 3.1.2.2). Future addition of genomic information on various yeast species with similar metabolism might help identification of bile acid pathway in strain POB-4.

Acknowledgment

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This study (Project No. 015675) was carried out with the support of National Institute of Agricultural Sciences, Rural Development Administration, Republic of Korea.

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