scholarly journals Mycoplasma hyopneumoniae p65 Surface Lipoprotein Is a Lipolytic Enzyme with a Preference for Shorter-Chain Fatty Acids

2004 ◽  
Vol 186 (17) ◽  
pp. 5790-5798 ◽  
Author(s):  
Jono A. Schmidt ◽  
Glenn F. Browning ◽  
Philip F. Markham
Keyword(s):  
Fatty Acids ◽  
Signal Sequence ◽  
Lipolytic Activity ◽  
Mycoplasma Hyopneumoniae ◽  
Lipolytic Enzyme ◽  
Protein Cleavage ◽  
Lipolytic Enzymes ◽  
Hydrolysis Of ◽  
Chain Fatty Acids

ABSTRACT Mycoplasma hyopneumoniae is the most significant bacterial pathogen of the respiratory tract of swine. p65 is an immunodominant surface lipoprotein of M. hyopneumoniae that is specifically recognized during disease. Analysis of the translated amino acid sequence of the gene encoding p65 revealed similarity to the GDSL family of lipolytic enzymes. To examine the lipolytic activity of p65, the gene was cloned and expressed in Escherichia coli after truncation of the prokaryotic lipoprotein signal sequence and mutagenesis of the mycoplasma TGA tryptophan codons. After treatment with thrombin, the recombinant glutathione S-transferase (GST)-p65 protein yielded a 66-kDa fusion protein cleavage product corresponding in size to the mature p65 protein. The esterase activity of recombinant GST-p65 was indicated by the formation of a cleared zone on tributyrin agar plates and the hydrolysis of p-nitrophenyl esters of caproate (pNPC) and p-nitrophenyl esters of palmitate (pNPP). Lipase activity was indicated by the hydrolysis of the artificial triglyceride 1,2-O-dilauryl-rac-glycero-3-glutaric acid resorufin ester. Using pNPC and pNPP as substrates, recombinant GST-p65 had optimal activity between pHs 9.2 and 10.2 and at a temperature higher than 39°C. Calcium ions did not increase the activity of recombinant GST-p65. Rabbit anti-p65 antibodies inhibited the activity of recombinant GST-p65 and also inhibited the growth of M. hyopneumoniae in vitro. Examination of the kinetic parameters of recombinant GST-p65 for the hydrolysis of pNPC and pNPP indicated a preference for the shorter fatty acid chain of pNPC. The physiological and/or pathogenic role of mycoplasma lipolytic enzymes has not been determined, but they are likely to play an important role in mycoplasmas' nutritional requirements for long-chain fatty acids and may reduce the function of lung surfactants in mycoplasma-induced respiratory diseases. This is the first report of the lipolytic activity of a lipid-modified surface immunogen of a mycoplasma.

scholarly journals Studies on lipid digestion in the preruminant calf. The source of lipolytic activity in the abomasum

1976 ◽  
Vol 36 (3) ◽  
pp. 439-447 ◽  
Author(s):  
Joyce Toothill ◽  
S. Y. Thompson ◽  
J. D. Edwards-Webb
Keyword(s):  
Fatty Acids ◽  
Dietary Fat ◽  
Milk Fat ◽  
Lipolytic Activity ◽  
Lipolytic Enzyme ◽  
Lipid Digestion ◽  
Lipolytic Enzymes ◽  
Ph Values ◽  
Proteolytic Activities ◽  
Pouch Secretions

1. Lipolytic and proteolytic activities and pH values were determined in secretions collected from innervated abomasal pouches and in abomasal contents from preruminant calves given liquid diets.2. No lipolytic activity was detected in pouch secretions collected during 1 h after feeding, though lipolytic activity was present in abomasal contents; pepsin (EC 3.4.23.1) and rennin (EC 3.4.23.4) were present in both pouch secretions and abomasal contents. The pH values of pouch secretions ranged from 1.2 to 1.8 and those of abomasal contents from 4.2 to 5.9.3. When diet was placed directly into the abomasal pouch soon after feeding, the pH values of pouch and abomasal contents decreased similarly (i.e. from 6.3 to approximately 5). Protease activity (U/ml) of pouch contents ranged from 0.1 to 0.8 and that of abomasal contents from 0.1 to 0.2. No lipolytic activity was detected in pouch contents, though abomasal contents contained 0.6 to 1.2 U/ml and when the diet contained milk-fat as the dietary fat source considerable lipolysis of triglycerides containing shorter-chain fatty acids was found.4. It is concluded that there is no significant secretion of lipolytic enzymes by the fundal mucosa and that the lipolysis of triglycerides in the abomasum of the preruminant calf is due predominantly to a lipolytic enzyme in saliva.

Short-Chain Fatty Acids Prevent Diabetic Nephropathy In Vivo and In Vitro

Diabetes ◽  
2018 ◽  
Vol 67 (Supplement 1) ◽  
pp. 92-OR ◽  
Author(s):  
WEI HUANG ◽  
YONG XU ◽  
YOUHUA XU ◽  
LUPING ZHOU ◽  
CHENLIN GAO
Keyword(s):  
Fatty Acids ◽  
Diabetic Nephropathy ◽  
Short Chain Fatty Acids ◽  
Short Chain ◽  
Chain Fatty Acids

PSIII-12 In vitro Evaluation of Purified Fiber Sources for Production of Short-chain Fatty Acids Using Pig Cecal Content as an Inoculum

2021 ◽  
Vol 99 (Supplement_1) ◽  
pp. 177-177
Author(s):  
Gabriela E Martinez Padilla ◽  
Rajesh Jha ◽  
Vivek Fellner ◽  
Eric van Heugten
Keyword(s):  
Fatty Acids ◽  
Enzymatic Hydrolysis ◽  
Resistant Starch ◽  
Potato Starch ◽  
Short Chain ◽  
Gas Liquid Chromatography ◽  
Citrus Peel ◽  
Starch Potato ◽  
Chain Fatty Acids

Abstract This study evaluated short-chain fatty acid (SCFA) production from purified fiber sources when fermented in vitro using pig cecal contents as an inoculum. Fiber sources of interest were inulin from chicory root (native and long-chain inulin with 90 and 98% fiber, respectively), pectin from citrus peel (high methoxyl pectin), resistant starch (native starch), potato starch (commercial grade), and β-glucan (β-1,3;β-1,6 yeast-derived). Cellulose and cornstarch were used as indigestible and highly digestible carbohydrates, respectively. Triplicate samples of substrates (2 g) were subjected to enzymatic hydrolysis with pepsin and pancreatin for 6 h. Subsequently, hydrolyzed residues (200 mg) were incubated under anaerobic conditions at 39°C with 30 mL solution of cecal inoculum collected from 3 sows fed a standard commercial diet and buffered mineral solution. After 48 h of incubation, solutions from fermented samples were analyzed for pH, SCFA, and branched-chain fatty acids (BCFA) using gas-liquid chromatography. Enzymatic hydrolysis had no effect on digestion of β-glucan, but total SCFA concentration after fermentation was highest (26.13 mmol/g) followed by resistant starch (22.61 mmol/g) and potato starch (22.20 mmol/g) and was lowest for cellulose (13.91 mmol/g). In contrast, native inulin was highly digested during enzymatic hydrolysis, resulting in the lowest substrate available for fermentation (11.84% DM) and the highest pH (5.98). Enzymatic hydrolysis and fermentation of resistant starch increased (P< 0.001) concentrations of acetate (0.60 mg/g), whereas potato starch and β-glucan yielded more butyrate (0.60 and 0.54 mg/g respectively), and β-glucan resulted in greater (P< 0.001) propionate concentrations (0.69 mg/g). Pectin resulted in the highest fermentation (82.38% DM disappearance) and the lowest pH (4.03) compared to the other fiber sources (P< 0.001) and yielded the lowest BCFA concentration (1.89 mM, P< 0.001). Results suggest that fermentation of resistant starch, potato starch, and β-glucan produced higher SCFA concentrations, while pectin resulted in a decreased pH of fermentation solution.

scholarly journals Characterization of the Gene Encoding the Major Secreted Lysophospholipase A of Legionella pneumophila and Its Role in Detoxification of Lysophosphatidylcholine

2002 ◽  
Vol 70 (11) ◽  
pp. 6094-6106 ◽  
Author(s):  
Antje Flieger ◽  
Birgid Neumeister ◽  
Nicholas P. Cianciotto
Keyword(s):  
Fatty Acids ◽  
Legionella Pneumophila ◽  
Lipolytic Activity ◽  
Cholesterol Acyltransferase ◽  
Phospholipase A ◽  
Wild Type ◽  
Gene Encoding ◽  
Acyltransferase Activity ◽  
Lipolytic Enzymes ◽  
Cloned Gene

ABSTRACT We previously showed that Legionella pneumophila secretes, via its type II secretion system, phospholipase A activities that are distinguished by their specificity for certain phospholipids. In this study, we identified and characterized plaA, a gene encoding a phospholipase A that cleaves fatty acids from lysophospholipids. The plaA gene encoded a 309-amino-acid protein (PlaA) which had homology to a group of lipolytic enzymes containing the catalytic signature GDSL. In Escherichia coli, the cloned gene conferred trypsin-resistant hydrolysis of lysophosphatidylcholine and lysophosphatidylglycerol. An L. pneumophila plaA mutant was generated by allelic exchange. Although the mutant grew normally in standard buffered yeast extract broth, its culture supernatants lost greater than 80% of their ability to release fatty acids from lysophosphatidylcholine and lysophosphatidylglycerol, implying that PlaA is the major secreted lysophospholipase A of L. pneumophila. The mutant's reduced lipolytic activity was confirmed by growth on egg yolk agar and thin layer chromatography and was complemented by reintroduction of an intact copy of plaA. Overexpression of plaA completely protected L. pneumophila from the toxic effects of lysophosphatidylcholine, suggesting a role for PlaA in bacterial detoxification of lysophospholipids. The plaA mutant grew like the wild type in U937 cell macrophages and Hartmannella vermiformis amoebae, indicating that PlaA is not essential for intracellular infection of L. pneumophila. In the course of characterizing plaA, we discovered that wild-type legionellae secrete a phospholipid cholesterol acyltransferase activity, highlighting the spectrum of lipolytic enzymes produced by L. pneumophila.

scholarly journals The impact of long-term dietary pattern of fecal donor on in vitro fecal fermentation properties of inulin

2016 ◽  
Vol 7 (4) ◽  
pp. 1805-1813 ◽  
Author(s):  
Junyi Yang ◽  
Devin J. Rose
Keyword(s):  
Fatty Acids ◽  
Short Chain Fatty Acids ◽  
Processed Meats ◽  
Branched Chain Fatty Acids ◽  
Fiber Plant ◽  
Branched Chain ◽  
The Impact ◽  
Chain Fatty Acids

A diet high in whole grains, dry beans, and certain vegetables that contributed dietary fiber, plant protein, and B vitamins resulted in high short chain fatty acids, while a diet high in diary and processed meats that provided cholesterol and little fiber resulted in high branched chain fatty acids and ammonia during fecal fermentation of inulin.

scholarly journals Profiles of Odd- and Branched-Chain Fatty Acids and Their Correlations With Rumen Fermentation Parameters, Microbial Protein Synthesis and Bacterial Populations Based on Pure Carbohydrate Incubation in Vitro

2020 ◽  
Author(s):  
Hangshu Xin ◽  
Xin Liu ◽  
Xin Jiang ◽  
Chunlong Liu ◽  
Shuzhi Zhang ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Volatile Fatty Acids ◽  
Cellulolytic Bacteria ◽  
Bacterial Populations ◽  
Branched Chain Fatty Acids ◽  
Degrading Bacteria ◽  
Branched Chain ◽  
Fermentation Parameters ◽  
Chain Fatty Acids

Abstract Background: The objectives of this study were to evaluate the profiles of odd- and branched-chain fatty acids (OBCFA; including C15:0, iso-C15:0, anteiso-C15:0, iso-C16:0, C17:0, iso-C17:0 and anteiso-C17:0) during pure carbohydrates incubation in vitro and whether they correlated with ruminal fermentation parameters, microbial crude protein (MCP) synthesis, and bacterial populations. The pure substrates containing five different ratios of fiber and starch (F:S; 0:100, 25:75, 50:50, 75:25 and 100:0) were incubated for 6 h, 12 h, 18 h and 24 h. Results: Except iso-C17:0, OBCFA concentrations were interacted by F:S and incubation time. The highest concentration of total OBCFA was found in the fermented mixture after 24 h of incubation when the F:S = 0:100; while the lowest level was 1.65 mg/g DM produced after 6 h of incubation with F:S = 50:50. The concentrations of total volatile fatty acids (TVFA) and MCP remarkably decreased linearly as the inclusion of fiber in the substrates increased, as expected. The proportions of investigated cellulolytic bacteria in our study were increased linearly (or linearly and quadratically) while those of R. amylophilus and S. bovis were decreased as fiber inclusion increased. The correlation analysis indicated that iso-C16:0 concentration might have potential as a marker of productions of TVFA and MCP with ρ being 0.78 and 0.82 respectively. Compared to starch degrading bacteria, cellulolytic bacteria had more correlations with OBCFA profiles, and the strongest association was found on the population of R. flavefaciens with C15:0 concentration (ρ = 0.70). Conclusions: Our study shows there might be scope for iso-C16:0 to predict rumen productions of VFA and MCP. Notedly, this is the first paper reporting linkage of OBCFA with rumen function based on pure carbohydrate in vitro incubation, which would avoid confounding interference from dietary protein and fat presence. However, more in-depth experiments are needed to substantiate the current findings.

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