Overexpression of nicotinamide mononucleotide adenylyltransferase (nmnat) increases the growth rate, Ca2+ concentration and cellulase production in Ganoderma lucidum

2020 ◽  
Vol 104 (16) ◽  
pp. 7079-7091
Author(s):  
Shengli Wang ◽  
Jing Han ◽  
Jiale Xia ◽  
Yanru Hu ◽  
Liang Shi ◽  
...  
2019 ◽  
Author(s):  
shengli Wang ◽  
jing Han ◽  
Jiale Xia ◽  
Yanru Hu ◽  
Liang Shi ◽  
...  

Abstract Background: There is an urgent need to search for new and economical ways to utilize diverse lignocellulose. Ganoderma lucidum is well-known edible medicinal fungus that has a strong ability to degrade a wide variety of cellulosic biomass and its nutrient utilization is closely related to extracellular cellulase. Nicotinamide adenine dinucleotide (NAD+), a nutritional sensor molecule, can respond to nutritional states and regulate cellular metabolism. Nicotinamide mononucleotide adenyltransferase (nmnat) is the key enzyme that catalyses the biosynthesis of NAD+.Result: In this study, a homologue of the gene encoding nmnat was cloned from G. lucidum. The Agrobacterium tumefaciens -mediated transformation (ATMT) method was used to construct the two overexpression strains OE:: nmnat4 and OE:: nmnat19 in G. lucidum. In the overexpression strains, the transcript levels of the nmnat gene and the NAD+ content were significantly increased. Gl nmnat overexpression strains showed dramatically stronger colony growth on different carbon sources, and the intracellular Ca 2+ concentration increased 3.95-fold and 2.10-fold in OE:: nmnat4 and OE:: nmnat19 , respectively, compared with WT strains. The CMCase activity increased by approximately 2.8-fold and 3-fold, and p NPGase activity increased by approximately 1.9-fold and 2.1-fold in OE:: nmnat4 and OE:: nmnat19 strains compared with the WT strains, respectively. Furthermore, it was found that NAD+ might induce cellulase production by regulating cytosolic Ca 2+ concentration.Conclusions: Taken together, our results revealed for the first time that NAD + could stimulate cellulase production and demonstrated that NAD + could increase the transcript levels of cellulase genes via the intracellular Ca 2+ concentration in G. lucidum. This research also provides a theoretical basis for conducting cellulase-related work on other basidiomycetes.


2020 ◽  
Author(s):  
Shengli Wang ◽  
Jing Han ◽  
Jiale Xia ◽  
Yanru Hu ◽  
Liang Shi ◽  
...  

Abstract Background: There is an urgent need to identify new and economical ways to utilize diverse types of lignocellulosic biomass. Ganoderma lucidum is well-known edible medicinal fungus that has an excellent ability to degrade a wide range of cellulosic biomass, and its nutrient utilization is closely related to the production of extracellular cellulase. Nicotinamide adenine dinucleotide (NAD+), a nutritional sensor molecule, can respond to nutritional states and regulate cellular metabolism, and nicotinamide mononucleotide adenyltransferase (nmnat) is the key enzyme that catalyses the biosynthesis of NAD+.Results: In this study, a homologue of the gene encoding nmnat was cloned from G. lucidum. The Agrobacterium tumefaciens-mediated transformation (ATMT) method was used to construct two G. lucidum overexpression strains, OE::nmnat4 and OE::nmnat19, in which the nmnat gene transcript levels and the NAD+ content were significantly increased. Glnmnat overexpression strains showed dramatically increased colony growth on different carbon sources, and the intracellular Ca2+ concentration was increased by 3.95- and 2.10-fold in the G. lucidum OE::nmnat4 and OE::nmnat19 strains, respectively, compared with that observed in the WT strain. The CMCase activity increased by approximately 2.8- and 3-fold, and that of pNPGase increased by approximately 1.9- and 2.1-fold in the G. lucidum OE::nmnat4 and OE::nmnat19 strains, respectively, compared with that observed in the WT strain. Furthermore, NAD+ was observed to potentially induce cellulase production by regulating the cytosolic Ca2+ concentration.Conclusions: Taken together, for the first time, our results revealed that NAD+ can stimulate cellulase production and increase the transcript levels of cellulase genes via increasing the intracellular Ca2+ concentration in G. lucidum. This research also provides a theoretical basis for conducting cellulase-related work in other basidiomycetes.


2019 ◽  
Vol 17 ◽  
Author(s):  
Farzane Kargar ◽  
Mojtaba Mortazavi ◽  
Mahmood Maleki ◽  
Masoud Torkzadeh Mahani ◽  
Younes Ghasemi ◽  
...  

Aims: The purpose of this study was to screen the bacteria producing cellulase enzymes and their bioinformatics studies. Background: Cellulose is a long-chain polymer of glucose that hydrolyzes by cellulases to glucose molecules. In order to design the new biotechnological applications, some strategies have been used as increasing the efficiency of enzyme production, generating cost-effective enzymes, producing stable enzymes and identification of new strains. Objective: On the other hand, some bacteria special features have made them suitable candidates for the identification of the new source of enzymes. In this regard, some native strains of bacteria were screened. Method: These bacteria were grown on a culture containing the liquid M9 media containing CMC to ensure the synthesis of cellulase. The formation of a clear area in the culture medium indicated decomposition of cellulose. In the following, the DNA of these bacteria were extracted and their 16S rDNA genes were amplified. Result: The results show that nine samples were able to synthesize cellulase. In following, these strains were identified using 16S rDNA. The results show that these screened bacteria belonged to the Bacillus sp., Alcaligenes sp., Alcaligenes sp., and Enterobacter sp.conclusionThe enzyme activity analysis shows that the Bacillus toyonensis, Bacillus sp. strain XA15-411 Bacillus cereus have produced the maximum yield of cellulases. However, these amounts of enzyme production in these samples are not proportional to their growth rate. As the bacterial growth chart within 4 consecutive days shows that the Alcaligenes sp. Bacillus cereus, Bacillus toyonensis, Bacillus sp. strain XA15-411 have a maximum growth rate. The study of the phylogenetic tree also shows that Bacillus species are more abundant in the production of cellulase enzyme. These bioinformatics analyses show that the Bacillus species have different evolutionary relationships and evolved in different evolutionary time. Other: However, for maximum cellulase production by this bacteria, some information as optimum temperature, optimum pH, carbon and nitrogen sources are needed for the ideal formulation of media composition. The cellulase production is closely controlled in microorganisms and the cellulase yields appear to depend on a variety of factors. However, the further studies are needed for cloning, purification and application of these new microbial cellulases in the different commercial fields as in food, detergent, and pharmaceutical, paper, textile industries and also various chemical industries. However, these novel enzymes can be further engineered through rational design or using random mutagenesis techniques.


Biochemistry ◽  
1986 ◽  
Vol 25 (12) ◽  
pp. 3725-3729 ◽  
Author(s):  
Paolo Natalini ◽  
Silverio Ruggieri ◽  
Nadia Raffaelli ◽  
Giulio Magni

2015 ◽  
Vol 110 (7) ◽  
pp. 890-897 ◽  
Author(s):  
Carlos H Niño ◽  
Nicolás Forero-Baena ◽  
Luis E Contreras ◽  
Diana Sánchez-Lancheros ◽  
Katherine Figarella ◽  
...  

2004 ◽  
Vol 11 (7) ◽  
pp. 873-885 ◽  
Author(s):  
G. Magni ◽  
A. Amici ◽  
M. Emanuelli ◽  
G. Orsomando ◽  
N. Raffaelli ◽  
...  

2013 ◽  
Vol 288 (29) ◽  
pp. 20908-20917 ◽  
Author(s):  
Tanjing Song ◽  
Leixiang Yang ◽  
Neha Kabra ◽  
Lihong Chen ◽  
John Koomen ◽  
...  

The chromosomal region encoding the nuclear NAD+ synthesis enzyme nicotinamide mononucleotide adenylyltransferase (NMNAT1) is frequently deleted in human cancer. We describe evidence that NMNAT1 interacts with the nucleolar repressor protein nucleomethylin and is involved in regulating rRNA transcription. NMNAT1 binds to nucleomethylin and is recruited into a ternary complex containing the NAD+-dependent deacetylase SirT1. NMNAT1 expression stimulates the deacetylase function of SirT1. Knockdown of NMNAT1 enhances rRNA transcription and promotes cell death after nutrient deprivation. Furthermore, NMNAT1 expression is induced by DNA damage and plays a role in preventing cell death after damage. Heterozygous deletion of NMNAT1 in lung tumor cell lines correlates with low expression level and increased sensitivity to DNA damage. These results suggest that NMNAT1 deletion in tumors may contribute to transformation by increasing rRNA synthesis, but may also increase sensitivity to nutrient stress and DNA damage.


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