Fenugreek galactomannan and citrus pectin improve several parameters associated with glucose metabolism and modulate gut microbiota in mice
. Nutrition 2018
, 134 - 142.e3. Publisher's VersionAbstract
ObjectiveGalactomannans derived from fenugreek confer known health benefits; however, there is little information regarding health benefits of citrus pectin (CP) and its association with gut microbiome metabolites. The aim of this study was to examine links between galactomannan and CP consumption, microbiota development, and glucose metabolism. Design Male C57 BL/6 J mice ages 7 to 8 wk were fed ad libitum with a normal diet or one supplemented with 15% of either galactomannan or CP. At 3 wk, an oral glucose tolerance test was performed. Animals were sacrificed at 4 wk and relevant organs were harvested. Results Fiber enrichment led to reductions in weight gain, fasting glucose levels, and total serum cholesterol (P < 0.05). Compared with mice fed the normal diet, microbiota populations were altered in both fiber groups and were found to be richer in Bacteroidetes rather than Firmicutes (P < 0.05). The modification was significantly greater in galactomannan-fed than in CP-fed mice (P < 0.0001). Also, enhanced levels of the short-chain fatty acid (SCFA) propionate were found in the cecal contents of CP-fed animals (P < 0.05). Protein expression levels of monocarboxylate transporter 1, which may promote transport of SCFA, were measured in the large intestines after fiber consumption. Enhanced adenosine monophosphate-activated protein kinase (AMPK) activation was observed in livers of galactomannan-fed mice (P < 0.05). Conclusion Consumption of diets containing soluble fibers, as used in this study, resulted in gut microbiota comprising a healthier flora, and led to positive effects on weight, glycemic control, and liver β oxidation via AMPK.
High Fat Diets Composed of Palm Stearin and Olive Oil Equally Exacerbate Liver Inflammatory Damage and Metabolic Stress in Mice
. Molecular Nutrition & Food Research 2018
, 1700915. Publisher's VersionAbstract
Scope People with fatty liver could be subject to acute infections such as sepsis. The aim of the study is to evaluate the effect of high fat diets (HFD) of olive oil and palm stearin on liver inflammation induced by lipopolysaccharides (LPS). Methods and results C57BL/6J male mice were treated with high fat diets with different sources of oils: palm stearin and olive oil for 8 weeks followed by LPS injection. The proinflammatory effect of olive oil was also studied using gavage treatment and IP injection of LPS. Animals fed with HFDs showed an increase in body weight, elevated blood glucose levels, and fatty liver phenotype. HFDs aggravated the effect of LPS treatment to induce inflammatory response compared to low fat diet (LFD) effect. Following HFD supplementation, LPS induced hyperinsulinemia, more liver damage than in animals that consumed LFD. In addition, both gavage and long-term feeding with high lipids in the presence of LPS resulted in inhibition of gluconeogenic genes expression. Conclusion HFDs of both monounsaturated and saturated fat potentiated liver inflammation induced by LPS treatment indicate that the total amount of fat consumed is the main proinflammatory factor rather than the type of fat.
Impact of Community-Based Nutrition Education on Geophagic Behavior and Dietary Knowledge and Practices among Rural Women in Nakuru Town, Kenya: A Pilot Study
. Journal of Nutrition Education and Behavior 2018
, 408 - 414.e1. Publisher's VersionAbstract
ObjectiveGeophagia, the deliberate consumption of rocks, soil, or clay, is prevalent in developing countries, particularly sub-Saharan Africa. Health risks associated with this behavior include parasitosis, heavy metal poisoning, nutrient deficiencies, and poor birth outcomes. This pilot study was designed to reduce geophagic practices and improve nutrition among rural Kenyan women. Methods The researchers used snowball sampling to recruit participants (n = 135; aged 15–49 years) from low socioeconomic areas who consumed geophagic materials. Interviews were carried out before and after a nutrition intervention implemented by trained community health volunteers. Results Nutrition education focusing on geophagia significantly (P < .001) decreased the practice in 77% of participants. Postintervention interviews also demonstrated substantial improvement in understanding the concept of making half the plate vegetables using the healthy plate model. Conclusions and Implications Nutrition education can be useful for reducing geophagia (a largely ignored, unsafe dietary behavior) and enhancing nutritional knowledge in African women.
“Practice What You Teach” Public Health Nurses Promoting Healthy Lifestyles (PHeeL-PHiNe): Program Evaluation
. The Journal of Ambulatory Care Management 2018
. Publisher's VersionAbstract
Healthy lifestyle programs are essential for meeting the challenge of noncommunicable diseases. The Public Health Nurses Promoting Healthy Lifestyles (PHeeL-PHiNe) program engaged nurses from family health clinics in Jerusalem District and included physical activity, healthy nutrition, and motivational skills. Questionnaires were completed at baseline, postintervention, and at 18 months. Results showed a marked effect on health practices. The proportion of nurses consuming a balanced diet and the use of food labels significantly increased and were maintained over time. Short-term improvements in physical activity were also observed. Nurses who practiced a healthy lifestyle were significantly more likely to provide guidance and counseling to families on healthy behaviors.
Effect of Pomegranate Juice on Intestinal Recovery Following Methotrexate-Induced Intestinal Damage in a Rat Model
. J Am Coll Nutr 2018
BACKGROUND/AIMS: Several studies have demonstrated the antimicrobial, antihelminthic, and antioxidant potential of the active ingredients of pomegranate (PMG) extracts, suggesting their preventive and curative role in several gastrointestinal disorders. In the present study, the authors evaluated the effects of oral PMG supplementation on intestinal structural changes, enterocyte proliferation, and apoptosis during methotrexate (MTX)-induced intestinal damage in a rat.
METHODS: Male rats were divided into 4 experimental groups: control rats; CONTR-PMG rats were treated with oral PMG given by gavage once a day 72 hours before and 72 hours following vehicle injection; MTX rats were treated with single dose of methotrexate; and MTX-PMG rats were treated with oral PMG following injection of MTX. Intestinal mucosal damage, mucosal structural changes, enterocyte proliferation, and enterocyte apoptosis were determined 72 hours following MTX injection. Western blotting was used to determine phosphorylated extracellular signal-regulated kinase (p-ERK) and caspase 3 protein levels.
RESULTS: MTX-PMG rats demonstrated greater jejunal and ileal bowel and mucosal weights, greater jejunal and ileal mucosal DNA and protein levels, greater villus height in jejunum and ileum and crypt depth in ileum, compared with MTX animals. A significant decrease in enterocyte apoptosis in ileum of MTX-PMG rats (vs MTX) was associated with a decrease in caspase 3 protein expression as well as increased cell proliferation, which was correlated with elevated p-ERK protein levels.
CONCLUSIONS: Treatment with oral PMG prevents mucosal injury and improves intestinal recovery following MTX injury in the rat.
Quercetin prevents small intestinal damage and enhances intestinal recovery during methotrexate-induced intestinal mucositis of rats
. Food Nutr Res 2018
Background: Gastrointestinal mucositis occurs as a consequence of cytotoxic treatment. Quercetin (QCT) is a bioflavonoid that exerts significant antioxidant activity and anti-inflammatory as well as anti-malignancy properties.
Objective: To evaluate the effects of oral QCT consumption in preventing intestinal mucosal damage and stimulating intestinal recovery following methotrexate (MTX)-induced intestinal damage in a rat model.
Design: Male Sprague-Dawley rats were divided into four groups: Control Group A (CONTR) - rats were treated with 2 cc of saline given by gavage for 6 days. Group B (CONTR-QCT) - rats were treated with QCT (100 mg/kg in 2 ml saline) given by gavage 3 days before and 3 days after intraperitoneal (IP) injection of saline. Group C (MTX) - rats were injected a single dose (25 mg/kg) of MTX IP. Group D (MTX-QCT) rats were treated with QCT (similar to Group B) 3 days before and 3 days after IP MTX injection. Intestinal mucosal parameters (bowel and mucosal weight, mucosal DNA and protein content, and villus height and crypt depth), enterocytes proliferation, and enterocyte apoptosis degree were investigated at sacrifice on the 4th day after MTX or saline injection.
Results: Administration of QCT to MTX-treated rats resulted in: (1) significant decrease in intestinal injury score, (2) significant increase in intestinal and mucosal weight in jejunum and ileum, (3) increase on the protein content of the ileum, (4) increase in the villus height in the ileum, (5) increase of crypt depth of jejunum and ileum, and (6) increase in cell proliferation in the jejunum and ileum compared to MTX-nontreated group.
Conclusions: Administration of QCT prevents intestinal damage and improves intestinal recovery following MTX-induced intestinal damage in a rat. We surmise that the effect of QCT is based on induction of cell proliferation in the crypt rather than inhibition of apoptosis.
Ostreolysin induces browning of adipocytes and ameliorates hepatic steatosis
. J Gastroenterol Hepatol 2018
BACKGROUND AND AIM: Non-alcoholic fatty liver disease (NAFLD) is associated with all features of the metabolic syndrome. Deposition of excess triglycerides in liver cells, a hallmark of NAFLD, is associated with loss of insulin sensitivity. Ostreolysin (Oly) is a 15-kDa fungal protein known to interact with cholesterol-enriched raft-like membrane domains. We aim to test whether a recombinant version of Oly (rOly) can induce functional changes in vitro in adipocytes or in vivo in mice fed a high-fat diet (HFD).
METHODS: White preadipocyte 3T3-L1 cells or mouse primary adipocytes treated with rOly. Male C57BL/6 mice were fed a control or HFD and treated with saline or with rOly (1 mg/kg BW) every other day for 4 weeks.
RESULTS: White preadipocyte 3T3-L1 cells or mouse primary adipocytes treated with rOly acquire a browning phenotype through activation of 5' adenosine monophosphate-activated protein kinase and downregulation of tumor necrosis factor α-mediated activation of IκB kinase ε and TANK-binding kinase 1. HFD-fed mice treated with rOly showed a 10% reduction in BW and improved glucose tolerance, which paralleled improved expression of liver and adipose functionality, metabolism, and inflammation status, mimicking the in vitro findings.
CONCLUSION: This study provides first evidence of rOly's prevention of HFD-induced NAFLD by stimulating liver and adipose muscle tissue functionality and oxidative potential, improving glucose tolerance, and ameliorating the metabolic profile of diet-induced obese mice.
Molecular Features Underlying Selectivity in Chicken Bitter Taste Receptors
. Frontiers in Molecular Biosciences 2018
6. Publisher's VersionAbstract
Chickens sense the bitter taste of structurally different molecules with merely three bitter taste receptors (Gallus gallus taste 2 receptors, ggTas2rs), representing a minimal case of bitter perception. Some bitter compounds like quinine, diphenidol and chlorpheniramine, activate all three ggTas2rs, while others selectively activate one or two of the receptors. We focus on bitter compounds with different selectivity profiles towards the three receptors, to shed light on the molecular recognition complexity in bitter taste. Using homology modeling and induced-fit docking simulations, we investigated the binding modes of ggTas2r agonists. Interestingly, promiscuous compounds are predicted to establish polar interactions with position 6.51 and hydrophobic interactions with positions 3.32 and 5.42 in all ggTas2rs; whereas certain residues are responsible for receptor selectivity. Lys3.29 and Asn3.36 are suggested as ggTas2r1-specificity-conferring residues; Gln6.55 as ggTas2r2-specificity-conferring residue; Ser5.38 and Gln7.42 as ggTas2r7-specificity conferring residues. The selectivity profile of quinine analogs, quinidine, epiquinidine and ethylhydrocupreine, was then characterized by combining calcium-imaging experiments and in-silico approaches. ggTas2r models were used to virtually screen BitterDB compounds. 50% of compounds known to be bitter to human are likely to be bitter to chicken, with 25%, 20%, 37% predicted to be ggTas2r1, ggTas2r2, ggTas2r7 agonists, respectively. Predicted ggTas2rs agonists can be tested with in-vitro and in-vivo experiments, contributing to our understanding of bitter taste in chicken and, consequently, to the improvement of chicken feed.
Independent Evolution of Strychnine Recognition by Bitter Taste Receptor Subtypes
. Frontiers in Molecular Biosciences 2018
9. Publisher's VersionAbstract
The 25 human bitter taste receptors (hT2Rs) recognize thousands of structurally and chemically diverse bitter substances. The binding modes of human bitter taste receptors hT2R10 and hT2R46, which are responsible for strychnine recognition, were previously established using site-directed mutagenesis, functional assays and molecular modeling. Here we construct a phylogenetic tree and reconstruct ancestral sequences of the T2R10 and T2R46 clades. We next analyze the binding sites in view of experimental data to predict their ability to recognize strychnine. This analysis suggests that the common ancestor of hT2R10 and hT2R46 is unlikely to bind strychnine in the same mode as either of its two descendants. Estimation of relative divergence times shows that hT2R10 evolved earlier than hT2R46. Strychnine recognition was likely acquired first by the earliest common ancestor of the T2R10 clade before the separation of primates from other mammals, and was highly conserved within the clade. It was probably independently acquired by the common ancestor of T2R43-47 before the homo-ape speciation, lost in most T2Rs within this clade, but enhanced in the hT2R46 after humans diverged from the rest of primates. Our findings suggest hypothetical strychnine T2R receptors in several species, and serve as an experimental guide for further study. Improved understanding of how bitter taste receptors acquire the ability to be activated by particular ligands is valuable for the development of sensors for bitterness and for potential toxicity.
The taste of KCl – What a difference a sugar makes
. Food Chemistry 2018
, 165 - 173. Publisher's VersionAbstract
Dramatic increase in NaCl consumption lead to sodium intake beyond health guidelines. KCl substitution helps reduce sodium intake but results in a bitter-metallic off-taste. Two disaccharides, trehalose and sucrose, were tested in order to untangle the chemical (increase in effective concentration of KCl due to sugar addition) from the sensory effects. The bitter-metallic taste of KCl was reduced by these sugars, while saltiness was enhanced or unaltered. The perceived sweetness of sugar, regardless of its type and concentration, was an important factor in KCl taste modulation. Though KCl was previously shown to increase the chemical activity of trehalose but not of sucrose, we found that it suppressed the perceived sweetness of both sugars. Therefore, sensory integration was the dominant factor in the tested KCl-sugar combinations.
Reengineering the ligand sensitivity of the broadly tuned human bitter taste receptor TAS2R14
. Biochimica et Biophysica Acta (BBA) - General Subjects 2018
, 2162 - 2173. Publisher's VersionAbstract
BackgroundIn humans, bitterness perception is mediated by ~25 bitter taste receptors present in the oral cavity. Among these receptors three, TAS2R10, TAS2R14 and TAS2R46, exhibit extraordinary wide agonist profiles and hence contribute disproportionally high to the perception of bitterness. Perhaps the most broadly tuned receptor is the TAS2R14, which may represent, because of its prominent expression in extraoral tissues, a receptor of particular importance for the physiological actions of bitter compounds beyond taste. Methods To investigate how the architecture and composition of the TAS2R14 binding pocket enables specific interactions with a complex array of chemically diverse bitter agonists, we carried out homology modeling and ligand docking experiments, subjected the receptor to point-mutagenesis of binding site residues and performed functional calcium mobilization assays. Results In total, 40 point-mutated receptor constructs were generated to investigate the contribution of 19 positions presumably located in the receptor's binding pocket to activation by 7 different TAS2R14 agonists. All investigated positions exhibited moderate to pronounced agonist selectivity. Conclusions Since numerous modifications of the TAS2R14 binding pocket resulted in improved responses to individual agonists, we conclude that this bitter taste receptor might represent a suitable template for the engineering of the agonist profile of a chemoreceptive receptor. General significance The detailed structure-function analysis of the highly promiscuous and widely expressed TAS2R14 suggests that this receptor must be considered as potentially frequent target for known and novel drugs including undesired off-effects.
BitterDB: taste ligands and receptors database in 2019
. Nucleic Acids Researchnar 2018
, D1179 - D1185. Publisher's VersionAbstract
) was introduced in 2012 as a central resource for information on bitter-tasting molecules and their receptors. The information in BitterDB is frequently used for choosing suitable ligands for experimental studies, for developing bitterness predictors, for analysis of receptors promiscuity and more. Here, we describe a major upgrade of the database, including significant increase in content as well as new features. BitterDB now holds over 1000 bitter molecules, up from the initial 550. When available, quantitative sensory data on bitterness intensity as well as toxicity information were added. For 270 molecules, at least one associated bitter taste receptor (T2R) is reported. The overall number of ligand–T2R associations is now close to 800. BitterDB was extended to several species: in addition to human, it now holds information on mouse, cat and chicken T2Rs, and the compounds that activate them. BitterDB now provides a unique platform for structure-based studies with high-quality homology models, known ligands, and for the human receptors also data from mutagenesis experiments, information on frequently occurring single nucleotide polymorphisms and links to expression levels in different tissues.
Molecular Features Underlying Selectivity in Chicken Bitter Taste Receptors
. Front Mol Biosci 2018
Chickens sense the bitter taste of structurally different molecules with merely three bitter taste receptors ( taste 2 receptors, ggTas2rs), representing a minimal case of bitter perception. Some bitter compounds like quinine, diphenidol and chlorpheniramine, activate all three ggTas2rs, while others selectively activate one or two of the receptors. We focus on bitter compounds with different selectivity profiles toward the three receptors, to shed light on the molecular recognition complexity in bitter taste. Using homology modeling and induced-fit docking simulations, we investigated the binding modes of ggTas2r agonists. Interestingly, promiscuous compounds are predicted to establish polar interactions with position 6.51 and hydrophobic interactions with positions 3.32 and 5.42 in all ggTas2rs; whereas certain residues are responsible for receptor selectivity. Lys and Asn are suggested as ggTas2r1-specificity-conferring residues; Gln as ggTas2r2-specificity-conferring residue; Ser and Gln as ggTas2r7-specificity conferring residues. The selectivity profile of quinine analogs, quinidine, epiquinidine and ethylhydrocupreine, was then characterized by combining calcium-imaging experiments and approaches. ggTas2r models were used to virtually screen BitterDB compounds. ~50% of compounds known to be bitter to human are likely to be bitter to chicken, with 25, 20, 37% predicted to be ggTas2r1, ggTas2r2, ggTas2r7 agonists, respectively. Predicted ggTas2rs agonists can be tested with and experiments, contributing to our understanding of bitter taste in chicken and, consequently, to the improvement of chicken feed.
RGS7 is recurrently mutated in melanoma and promotes migration and invasion of human cancer cells
. Scientific Reports 2018
653. Publisher's VersionAbstract
Analysis of 501 melanoma exomes revealed RGS7, which encodes a GTPase-accelerating protein (GAP), to be a tumor-suppressor gene. RGS7 was mutated in 11% of melanomas and was found to harbor three recurrent mutations (p.R44C, p.E383K and p.R416Q). Structural modeling of the most common recurrent mutation of the three (p.R44C) predicted that it destabilizes the protein due to the loss of an H-bond and salt bridge network between the mutated position and the serine and aspartic acid residues at positions 58 as 61, respectively. We experimentally confirmed this prediction showing that the p.R44C mutant protein is indeed destabilized. We further show RGS7 p.R44C has weaker catalytic activity for its substrate Gαo, thus providing a dual mechanism for its loss of function. Both of these effects are expected to contribute to loss of function of RGS7 resulting in increased anchorage-independent growth, migration and invasion of melanoma cells. By mutating position 56 in the R44C mutant from valine to cysteine, thereby enabling the formation of a disulfide bridge between the two mutated positions, we slightly increased the catalytic activity and reinstated protein stability, leading to the rescue of RGS7′s function as a tumor suppressor. Our findings identify RGS7 as a novel melanoma driver and point to the clinical relevance of using strategies to stabilize the protein and, thereby, restore its function.
Novel reagents for human prolactin research: large-scale preparation and characterization of prolactin receptor extracellular domain, non-pegylated and pegylated prolactin and prolactin receptor antagonist
. Protein Eng Des Sel 2018
To provide new tools for in vitro and in vivo prolactin (PRL) research, novel protocols for large-scale preparation of untagged human PRL (hPRL), a hPRL antagonist (del 1-9-G129R hPRL) that acts as a pure antagonist of hPRL in binding to hPRL receptor extracellular domain (hPRLR-ECD), and hPRLR-ECD are demonstrated. The interaction of del 1-9-G129R hPRL with hPRLR-ECD was demonstrated by competitive non-radioactive binding assay using biotinylated hPRL as the ligand and hPRLR-ECD as the receptor, by formation of stable 1:1 complexes with hPRLR-ECD under non-denaturing conditions using size-exclusion chromatography, and by surface plasmon resonance methodology. In all three types of experiments, the interaction of del 1-9-G129R hPRL was equal to that of unmodified hPRL. Del 1-9-G129R hPRL inhibited the hPRL-induced proliferation of Baf/LP cells stably expressing hPRLR. Overall, the biological properties of del 1-9-G129R hPRL prepared by the protocol described herein were similar to those of the antagonist prepared using the protocol reported in the original study; however, the newly described protocol improved yields by >6-fold. To provide long-lasting hPRL as a new reagent needed for in vivo experiments, we prepared its mono-pegylated analogue and found that pegylation lowers its biological activity in a homologous in vitro assay. As its future use will require the development of a PRL antagonist with highly elevated affinity, del 1-9-G129R hPRL was expressed on the surface of yeast cells. It retained its binding capacity for hPRLR-ECD, and this methodology was shown to be suitable for future development of high-affinity hPRL antagonists using a library of randomly mutated open reading frame of del 1-9-G129R hPRL and selecting high-affinity mutants by yeast surface display methodology.
Inactivation kinetics for Salmonella typhimurium in red pepper powders treated by radio frequency heating
. Food Control 2018
, 437 - 442. Publisher's VersionAbstract
Radio frequency (RF) dielectric heating has been investigated to inactivate pathogens in some low-moisture foods. This study was aimed to evaluate RF inactivation effects on Salmonella typhimurium in red pepper powders, by focusing on the influence of sample initial water activity (aw) and applying Weibull model to describe the inactivation curves. The experimental results showed that RF heating rate increased when aw was in the range of 0.57–0.71, but decreased after aw reached to 0.71. During the come-up time of RF heating, 2–3 log reduction of the pathogen was achieved depending on aw levels. Increasing initial aw could first increased log reductions and then decreased the log reductions, optimum aw level was 0.71 for RF inactivation of Salmonella in red pepper powders. For red pepper powders with aw of 0.71, RF heating to 70 °C (come-up time was 110 s) with holding time over 60 s could achieve >5 log reduction of S. typhimurium. Weibull model well fitted the survival curves of the pathogen with goodness of fit (R2 > 0.93, RMSE<0.29). Scale factor (b) of the model increased with treatment temperature increasing, while the shape factor (n) was independent on temperature. This study provided basic guideline for using RF heating to inactivate Salmonella in red pepper powders.
Random peptide mixtures as new crop protection agents
. Microb Biotechnol 2018
Many types of crops are severely affected by at least one important bacterial disease. Chemical control of bacterial plant diseases in the field vastly relies on copper-based bactericides, yet with limited efficacy. In this study, we explored the potential of two random peptide mixture (RPM) models as novel crop protection agents. These unique peptide mixtures consist of random combination of l-phenylalanine and l- or d-lysine (FK-20 and FdK-20, respectively) along the 20 mer chain length of the peptides. Both RPMs displayed powerful bacteriostatic and bactericidal activities towards strains belonging to several plant pathogenic bacterial genera, for example, Xanthomonas, Clavibacter and Pseudomonas. In planta studies in the glasshouse revealed that RPMs significantly reduced disease severity of tomato and kohlrabi plants infected with Xanthomonas perforans and Xanthomonas campestris pv. campestris respectively. Moreover, RPM effects on reduction in disease severity were similar to those exerted by the commercial copper-based bactericide Kocide 2000 that was applied at a 12-fold higher concentration of the active compound relative to the RPM treatments. Importantly, the two tested RPM compounds had no toxic effect on survival of bees and Caco-2 mammalian cells. This study demonstrates the potential of these innovative RPMs to serve as crop protection agents against crop diseases caused by phytopathogenic bacteria.
Random mixtures of antimicrobial peptides inhibit bacteria associated with pasteurized bovine milk
. J Pept Sci 2018
The shelf life of pasteurized bovine milk is limited by microorganism activity as surviving bacteria continue to grow in the bovine milk, eventually causing milk spoilage. In the current study, we used matrix-assisted laser desorption ionization time of flight mass spectrometry to identify pasteurized bovine milk-associated mesophilic and psychrotrophic bacteria. We have recently designed random cationic peptide mixtures that possess strong antimicrobial and antibiofilm properties. These compounds are cheap and easy to synthesize and represent a new class of antimicrobial agents. Here, we show that the random peptide mixtures are able to efficiently eradicate the bacteria identified as associated with pasteurized bovine milk, and reduced significantly the growth of Bacillus subtilis in milk. We propose these antimicrobial peptides as potential candidates for integration in bioactive milk and food packaging to prevent bacterial growth and extend the shelf life of food.
Preparation of biologically active monomeric recombinant zebrafish (Danio rerio) and rainbow trout (Oncorhynchus mykiss) recombinant growth hormones
. Fish Physiology and Biochemistry 2018
, 1215 - 1222. Publisher's VersionAbstract
Fish growth hormones (GHs) play an important role in regulating growth, metabolism, reproduction, osmoregulation, and immunity and have thus garnered attention for their application in aquaculture. Zebrafish GH (zGH) cDNA or rainbow trout GH (rtGH) cDNA was cloned into the pMon3401 vector, expressed in MON105-competent Escherichia coli and purified to homogeneity. Their biological activity was evidenced by their ability to interact with ovine GH receptor extracellular domain and stimulate GH receptor-mediated proliferation in FDC-P1-3B9 cells stably transfected with rabbit GH receptor. The relative affinity of zGH and rtGH, estimated by IC50, was about 38-fold and 512-fold lower, respectively, than ovine GH. This is likely the reason for the low biological activity in cells with rabbit GH receptor, ~ 36-fold lower for zGH and ~ 107-fold lower for rtGH than for human GH. This was not due to improper refolding, as evidenced by circular dichroism analysis. Predicting the activity of fish GHs is problematic as there is no one single optimal in vitro bioassay; heterologous assays may be ambiguous, and only homologous assays are suitable for measuring activity.