Dataset Information

Molecular cloning, purification, expression, and characterization of ?-1, 4-endoglucanase gene (Cel5A) from Eubacterium cellulosolvens sp. isolated from Holstein steers' rumen.

ABSTRACT: OBJECTIVE:This study was conducted to isolate the cellulolytic microorganism from the rumen of Holstein steers and characterize endoglucanase gene (Cel5A) from the isolated microorganism. METHODS:To isolate anaerobic microbes having endoglucanase, rumen fluid was obtained from Holstein steers fed roughage diet. The isolated anaerobic bacteria had 98% similarity with Eubacterium cellulosolvens (E. cellulosolvens) Ce2 (Accession number: AB163733). The Cel5A from isolated E. cellulolsovens sp. was cloned using the published genome sequence and expressed through the Escherichia coli BL21. RESULTS:The maximum activity of recombinant Cel5A (rCel5A) was observed at 50°C and pH 4.0. The enzyme was constant at the temperature range of 20°C to 40°C but also, at the pH range of 3 to 9. The metal ions including Ca2+, K+, Ni2+, Mg2+, and Fe2+ increased the endoglucanase activity but the addition of Mn2+, Cu2+, and Zn2+ decreased. The Km and Vmax value of rCel5A were 14.05 mg/mL and 45.66 ?mol/min/mg. Turnover number, Kcat and catalytic efficiency, Kcat/Km values of rCel5A was 96.69 (s-1) and 6.88 (mL/mg/s), respectively. CONCLUSION:Our results indicated that rCel5A of E. cellulosolvens isolated from Holstein steers had a broad pH range with high stability under various conditions, which might be one of the beneficial characteristics of this enzyme for possible industrial application.

SUBMITTER: Park T

PROVIDER: S-EPMC5838334 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Molecular cloning, purification, expression, and characterization of β-1, 4-endoglucanase gene (<i>Cel5A</i>) from <i>Eubacterium cellulosolvens</i> sp. isolated from Holstein steers' rumen.

Park Tansol T Seo Seongwon S Shin Teaksoon T Cho Byung-Wook BW Cho Seongkeun S Kim Byeongwoo B Lee Seyoung S Ha Jong K JK Seo Jakyeom J

Asian-Australasian journal of animal sciences. 20170918 4

<h4>Objective</h4>This study was conducted to isolate the cellulolytic microorganism from the rumen of Holstein steers and characterize endoglucanase gene (<i>Cel5A</i>) from the isolated microorganism.<h4>Methods</h4>To isolate anaerobic microbes having endoglucanase, rumen fluid was obtained from Holstein steers fed roughage diet. The isolated anaerobic bacteria had 98% similarity with <i>Eubacterium cellulosolvens</i> (<i>E. cellulosolvens</i>) Ce2 (Accession number: AB163733). The <i>Cel5A</ ...[more]

PMID: 28920412

Similar Datasets

Project description:Seasonal effects on rumen microbiome and enteric methane (CH4) emissions are poorly documented. In this study, 6 Holstein and 6 Jersey steers were fed the same total mixed ration diet during winter, spring, and summer seasons under a 2 × 3 factorial arrangement for 30 days per season. The dry matter intake (DMI), rumen fermentation characteristics, enteric CH4 emissions and rumen microbiota were analyzed. Holstein had higher total DMI than Jersey steers regardless of season. However, Holstein steers had the lowest metabolic DMI during summer, while Jersey steers had the lowest total DMI during winter. Jersey steers had higher CH4 yields and intensities than Holstein steers regardless of season. The pH was decreased, while ammonia nitrogen concentration was increased in summer regardless of breed. Total volatile fatty acids concentration and propionate proportions were the highest in winter, while acetate and butyrate proportion were the highest in spring and in summer, respectively, regardless of breed. Moreover, Holstein steers produced a higher proportion of propionate, while Jersey steers produced a higher proportion of butyrate regardless of season. Metataxonomic analysis of rumen microbiota showed that operational taxonomic units and Chao 1 estimates were lower and highly unstable during summer, while winter had the lowest Shannon diversity. Beta diversity analysis suggested that the overall rumen microbiota was shifted according to seasonal changes in both breeds. In winter, the rumen microbiota was dominated by Carnobacterium jeotgali and Ruminococcus bromii, while in summer, Paludibacter propionicigenes was predominant. In Jersey steers, Capnocytophaga cynodegmi, Barnesiella viscericola and Flintibacter butyricus were predominant, whereas in Holstein steers, Succinivibrio dextrinosolvens and Gilliamella bombicola were predominant. Overall results suggest that seasonal changes alter rumen microbiota and fermentation characteristics of both breeds; however, CH4 emissions from steers were significantly influenced by breeds, not by seasons.

Project description:Previous studies have focused on the rumen microbiome and enteric methane (CH4) emissions in dairy cows, yet little is known about steers, especially steers of dairy breeds. In the present study, we comparatively examined the rumen microbiota, fermentation characteristics, and CH4 emissions from six non-cannulated Holstein (710.33 ± 43.02 kg) and six Jersey (559.67 ± 32.72 kg) steers. The steers were fed the same total mixed ration (TMR) for 30 days. After 25 days of adaptation to the diet, CH4 emissions were measured using GreenFeed for three consecutive days, and rumen fluid samples were collected on last day using stomach tubing before feeding (0 h) and 6 h after feeding. CH4 production (g/d/animal), CH4 yield (g/kg DMI), and CH4 intensity (g/kg BW0.75) were higher in the Jersey steers than in the Holstein steers. The lowest pH value was recorded at 6 h after feeding. The Jersey steers had lower rumen pH and a higher concentration of ammonia-nitrogen (NH3-N). The Jersey steers had a numerically higher molar proportion of acetate than the Holstein steers, but the opposite was true for that of propionate. Metataxonomic analysis of the rumen microbiota showed that the two breeds had similar species richness, Shannon, and inverse Simpson diversity indexes. Principal coordinates analysis showed that the overall rumen microbiota was different between the two breeds. Both breeds were dominated by Prevotella ruminicola, and its highest relative abundance was observed 6 h after feeding. The genera Ethanoligenens, Succinivibrio, and the species Ethanoligenens harbinense, Succinivibrio dextrinosolvens, Prevotella micans, Prevotella copri, Prevotella oris, Prevotella baroniae, and Treponema succinifaciens were more abundant in Holstein steers while the genera Capnocytophaga, Lachnoclostridium, Barnesiella, Oscillibacter, Galbibacter, and the species Capnocytophaga cynodegmi, Galbibacter mesophilus, Barnesiella intestinihominis, Prevotella shahii, and Oscillibacter ruminantium in the Jersey steers. The Jersey steers were dominated by Methanobrevibacter millerae while the Holstein steers by Methanobrevibacter olleyae. The overall results suggest that sampling hour has little influence on the rumen microbiota; however, breeds of steers can affect the assemblage of the rumen microbiota and different mitigation strategies may be needed to effectively manipulate the rumen microbiota and mitigate enteric CH4 emissions from these steers.

Dataset Information

Molecular cloning, purification, expression, and characterization of ?-1, 4-endoglucanase gene (Cel5A) from Eubacterium cellulosolvens sp. isolated from Holstein steers' rumen.

Publications

Molecular cloning, purification, expression, and characterization of β-1, 4-endoglucanase gene (<i>Cel5A</i>) from <i>Eubacterium cellulosolvens</i> sp. isolated from Holstein steers' rumen.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets