In Silico Structural and Functional Analysis of the Mitochondrial Malate Transporters in Oleaginous Fungus Mucor circinelloides WJ11
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Abstract:
Malate transporter proteins (MTPs) play a pivotal role in regulating flux in the citrate/malate/pyruvate shuttle to deliver acetyl-CoA from the mitochondria to the cytosol and thus regulate lipid biosynthesis in oleaginous fungi. Despite the recent successful exploration of the mitochondrial malate transporters in Mucor circinelloides, research with in silico analyses that include molecular docking and their dynamics, in addition to homology modelling of malate transporters, have not been reported. In this study, the physico-chemical properties and nucleotide sequence analysis of two mitochondrial MTPs (MT and SoDIT-a with Gene/protein ID scafold00018.48 and scafold00239.15, respectively), in M. circinelloides WJ11 were performed. The three-dimensional (3D) model of the mitochondrial MTPs was determined and the best-docked complex stabilities were demonstrated with molecular dynamic (MD) simulations. The activity domain was revealed to form hydrogen bonds and piling interactions with citrate and malate upon docking. Our study showed better binding affinities for the MTPs—reaching up to −3.44 and −7.27 kcal/mol with the MT and SoDIT-a proteins, respectively (compared to the target of −2.85 and −6.00 kcal/mol for citric acid-binding). MD simulations illustrated that the protein complexes demonstrated conformational stability throughout the simulation. This study was the first to elucidate the structural characteristics of mitochondrial MTPs in M. circinelloides WJ11, providing direct evidence regarding the transport mechanism of specific substrates. Furthermore, the current results support ongoing efforts to combine functional and structural data to better understand the MTPs (at the molecular and atomic levels) of an oleaginous fungus such as M. circinelloides.Keywords:
Mucor circinelloides
Malate dehydrogenase
The effects of various carbon sources and cultivation conditions on the growth kinetics, lipid accumulation, and medium-chain fatty acid (MCFA) production of Mucor circinelloides (MC) was investigated for 72 h in shake flask cultivation. Our previous investigation reported increments of 28 to 46% MCFAs among total cell lipids when the MC genome was genetically modified, in comparison to the wild-type. However, the growth of the engineered strain M65-TE-04 was adversely affected. Therefore, the current study was designed to enhance the growth, lipid production, and MCFA productivity of engineered M. circinelloides by optimizing the pH, agitation speed, temperature, and carbon sources. The findings for individual variables disclosed that the highest biomass (17.0 g/L) was obtained when coconut oil mixed with glucose was used as a carbon source under normal culture conditions. Additionally, the maximum lipid contents (67.5% cell dry weight (CDW)), MCFA contents (53% total fatty acid (TFA)), and overall lipid productivity (3.53 g/L·d) were attained at 26 °C, pH 6.0, and 150 rpm, respectively. The maximum biomass (19.4 g/L), TFA (14.3g/L), and MCFA (4.71 g/L) contents were achieved with integration of a temperature of 26 °C, pH 6.0, agitation speed 300 rpm, and coconut oil mixed medium as the carbon source. This work illustrates that biomass, TFA, and MCFA contents were increased 1.70–2.0-fold by optimizing the initial pH, agitation speed, temperature, and carbon sources in the M. circinelloides engineered strain (M65-TE-04) in comparison to initial cultivation conditions.
Mucor circinelloides
Carbon fibers
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Annals of the XIII International Fungal Biology Conference (IFBC) & IV International Symposium on Fungal Stress (ISFUS) (978-85-5722-449-0) - Effect Of Inhibitory Phenolic Compounds Over The Carotenoids Production By The Oleaginous Fungi Mucor Circinelloides
Mucor circinelloides
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Mucor circinelloides
Malic enzyme
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Mucor species are common soil fungi but also known as agents of human infections (mucormycosis) and used in food production and biotechnology. Mucor circinelloides is the Mucor species that is most frequently isolated from clinical sources. The taxonomy of Mucor circinelloides and its close relatives (Mucor circinelloides complex - MCC) is still based on morphology and mating behaviour. The aim of the present study was a revised taxonomy of the MCC using a polyphasic approach. Using a set of 100 strains molecular phylogenetic analysis of five markers (ITS, rpb1, tsr1, mcm7, and cfs, introduced here) were performed, combined with phenotypic studies, mating tests and the determination of the maximum growth temperatures. The multi-locus analyses revealed 16 phylogenetic species of which 14 showed distinct phenotypical traits and were recognised as discrete species. Five of these species are introduced as novel taxa: M. amethystinus sp. nov., M. atramentarius sp. nov., M. variicolumellatus sp. nov., M. pseudocircinelloides sp. nov., and M. pseudolusitanicus sp. nov. The former formae of M. circinelloides represent one or two separate species. In the MCC, the simple presence of well-shaped zygospores only indicates a close relation of both strains, but not necessarily conspecificity. Seven species of the MCC have been implemented in human infection: M. circinelloides, M. griseocyanus, M. janssenii, M. lusitanicus, M. ramosissimus, M. variicolumellatus, and M. velutinosus.
Mucor circinelloides
Species complex
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Mucor circinelloides
Malic enzyme
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Objective:To study the influence of carbon sources onγ-linolenic acid production by Mucor circinelloides EIM-10 and make a solid foundation for regulating GLA producing of Mucor circinelloides EIM-10.Method: Dry cell weight and lipid yield of mucor circinelloides and GLA of the oil content was determinated.Result:While glucose was used as carbon source,only 2% lipid of its dry cell weight which contained 18%γ-linolenic acid was accumulated by Mucor circinelloides EIM-10;While soybean oil was used as carbon source,the biomass of Mucor circinelloides was 33g/L and the lipid content was 35% of its dry cell weight with a GLA content of 3%.Acetic acid and citric acid could not be utilized but sodium acetate and sodium citrate could be utilized without lipid accumulation by Mucor circinelloides EIM-10.Conclusion:Ectogenesis fatty acid could be utlized to accumulate lipid but little fatty acid could be synthesized to produce oil by the strain Mucor circinelloides EIM-10.
Mucor circinelloides
gamma-Linolenic acid
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Исследовали рост, липогенез, морфологию и активность ферментов цикла трикарбоновых кислот и глиоксилатного цикла гриба Mucor circinelloides var. lusitanicus ИНМИ при различных концентрациях в среде источника азота (мочевины) и способах его введения. Показано, что максимальное количество липидов в биомассе накапливалось при низкой концентрации источника азота (0.5 г/л), в то время как максимальное содержание -линоленовой кислоты в липидах при высокой (до 4.0 г/л). Установлено, что при внесении источника азота в режиме с подпиткой, по сравнению с тем же его количеством, внесенным однократно, содержание -линоленовой кислоты в сумме жирных кислот возрастало до 35%. Отмечены различия в составе жирных кислот и степени ненасыщенности липидов различных субклеточных фракций. В культуре, выращенной при однократной подаче источника азота, наблюдалась выраженная деформация мицелия, при этом активность ферментов ЦТК НАД-зависимой изоцитратдегидрогеназы (ИЦДГ) и малатдегидрогеназы (МДГ) была ниже, чем в варианте с подпиткой, а активность ключевого фермента глиоксилатного цикла изоцитратлиазы (ИЦЛ) выше. Обсуждается связь особенностей метаболических процессов и состава липидов с дифференцировкой гриба в зависимости от режима азотного питания.
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Mucor circinelloides
Methyl jasmonate
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Mucor circinelloides is a fungus that belongs to the order Mucorales. It grows as mold in the environment and can cause mucormycosis, a potentially fatal infection in immunocompromised patients. M. circinelloides is a biodiesel producer and serves as a model organism for studying several biological processes, such as light responses and RNA interference-mediated gene silencing. Over the past decade, the increasing number of molecular tools has also allowed us to manipulate the genome of this fungus. This article outlines the fundamental protocols for the in vitro growth, maintenance, and genetic manipulation of M. circinelloides in the laboratory. © 2018 by John Wiley & Sons, Inc.
Mucor circinelloides
Mucorales
Filamentous fungus
Model Organism
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Mucor circinelloides
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