Mechanisms of Dipeptide Uptake by Rat Small Intestine in Vitro

1971 ◽  
Vol 40 (3) ◽  
pp. 247-259 ◽  
Author(s):  
B. Cheng ◽  
F. Navab ◽  
M. T. Lis ◽  
T. N. Miller ◽  
D. M. Matthews
Keyword(s):  
Amino Acids ◽  
Small Intestine ◽  
Amino Acid ◽  
Acid Uptake ◽  
Rat Small Intestine ◽  
Amino Acid Oxidase ◽  
Wide Range ◽  
Mucosal Cells ◽  
Hydrolysis Of

1. The uptake of l-methionine and glycine as free amino acids, and from their dipeptides by everted rings of rat small intestine in vitro has been investigated. The concentrations used covered a wide range, including values likely to be near those found in the lumen of the intestine. 2. Though no intact peptides were found in the mucosal cells, evidence was obtained which showed that hydrolysis of the peptides was cellular at all concentrations. Total hydrolysis of peptides by the intestine was very great in relation to amino acid uptake over very short incubation times, suggesting that much hydrolysis took place superficially. 3. Except at the lowest concentrations, the rates of uptake of amino acids from the peptides were more rapid than from the equivalent amino acid mixtures. Competition for uptake between glycine and methionine was avoided when they were presented in the form of l-methionylglycine. 4. Anoxia inhibited uptake of methionine from free l-methionine and from l-methionyl-l-methionine. It also inhibited hydrolysis of l-methionyl-l-methionine by intact intestine, but not by intestinal homogenates, suggesting that peptide uptake may be energy-dependent. The l-amino acid oxidase of snake venom, which destroys l-methionine but has no effect on glycine or on the peptides studied, inhibited methionine uptake from peptides when present at high concentrations, suggesting that a major site of hydrolysis is enzyme-accessible. 5. It is suggested that there may be two modes of uptake of amino acids from oligopeptides: (1) surface hydrolysis by mechanisms closely linked to the amino acid entry mechanisms, and (2) peptide entry into the mucosal cells by a special mechanism, followed by intracellular hydrolysis.

scholarly journals Action of growth hormone in vitro on the net uptake and incorporation into protein of amino acids in muscle from intact rabbits given protein-deficient diets

1976 ◽  
Vol 35 (1) ◽  
pp. 1-10 ◽  
Author(s):  
M. R. Turner ◽  
P. J. Reeds ◽  
K. A. Munday
Keyword(s):  
Amino Acids ◽  
Growth Hormone ◽  
Protein Synthesis ◽  
Amino Acid ◽  
De Novo ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
Amino Acid Incorporation ◽  
Acid Incorporation

1. Net amino acid uptake, and incorporation into protein have been measured in vitro in the presence and absence of porcine growth hormone (GH) in muscle from intact rabbits fed for 5 d on low-protein (LP), protein-free (PF) or control diets.2. In muscle from control and LP animals GH had no effect on the net amino acid uptake but stimulated amino acid incorporation into protein, although this response was less in LP animals than in control animals.3. In muscle from PF animals, GH stimulated both amino acid incorporation into protein and the net amino acid uptake, a type of response which also occurs in hypophysectomized animals. The magnitude of the effect of GH on the incorporation of amino acids into protein was reduced in muscle from PF animals.4. The effect of GH on the net amino acid uptake in PF animals was completely blocked by cycloheximide; the uptake effect of GH in these animals was dependent therefore on de novo protein synthesis.5. It is proposed that in the adult the role of growth hormone in protein metabolism is to sustain cellular protein synthesis when there is a decrease in the level of substrate amino acids, similar to that which occurs during a short-term fast or when the dietary protein intake is inadequate.

An Automated Detection System for Most L-α-Amino Acids

1969 ◽  
Vol 15 (2) ◽  
pp. 154-161 ◽  
Author(s):  
K Van Dyke ◽  
C Szustkiewicz
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Detection System ◽  
Automated System ◽  
Acid Oxidase ◽  
Enzymatic Reactions ◽  
Amino Acid Oxidase ◽  
Automated Method ◽  
Wide Range

Abstract An automated system for the determination of the L-α form of the majority of amino acids is presented. The method is based upon oxidative deamination of the amino acid coupled with oxidation of o-dianisidine by hydrogen peroxide. This procedure can be used comparatively for the determination of a mixture of L-α-amino acids or for the majority of separated L-α-amino acids (especially in conjunction with column separations from urine and blood which give falsely positive identification with ninhydrin detection). The stereospecific nature of the L-α-amino acid oxidase enables the investigator to quantitate the amount of L-α-amino acid in the presence of the D-α form. From an academic viewpoint, the extreme sensitivity and wide range of the detection system make it advantageous for the study of the enzyme itself. This automated method also may be employed to follow enzymatic reactions—e.g., those catalyzed by peptidases or racemases. The methodology is extremely convenient with good reagent stability and is much more sensitive than manometric technics.

In Vitro Measurement of Amino-Acid Uptake by the Human Small Intestine

Pharmacology ◽  
1964 ◽  
Vol 10 (3) ◽  
pp. 157-163 ◽  
Author(s):  
J.W.L. Robinson ◽  
F. Taminelli ◽  
J.P. Felber ◽  
P. Magnenat
Keyword(s):  
Small Intestine ◽  
Amino Acid ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
Human Small Intestine

Trans-tegumental absorption of L-alanine and L-leucine by a monogenean, Diclidophora merlangi

Parasitology ◽  
1978 ◽  
Vol 76 (1) ◽  
pp. 29-37 ◽  
Author(s):  
D. W. Halton
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Amino Acid ◽  
Sea Water ◽  
Low Molecular Weight ◽  
Acid Uptake ◽  
Neutral Amino Acids ◽  
Freeze Dried ◽  
Significant Difference

SummaryAn in vitro investigation has been made of the relative roles of the gut and tegument in the absorption of the neutral amino acids L-alanine and L-leucine by a marine fish-gill parasite, Diclidophora merlangi. The use of ligatures to preclude oral ingestion of trace-labelled medium has proved inadequate, invariably damaging the tegument, as revealed by stereoscan electron microscopy, and resulting in artifactual levels of absorption. Three alternative procedures have given consistently reliable data on the route of entry of low molecular weight substrates. (1) Ultrastructural examination of worms previously incubated in electron-dense cationic tracers has shown that, in vitro, there is no oral intake of sea water. (2) The suspending of worms in trace-labelled medium with the mouth out of the medium and comparing amino acid uptake with that of worms totally immersed in medium has revealed no statistically significant difference in the absorption levels. (3) Application of section (freeze-dried) auto-radiography to detect diffusible isotope has demonstrated directly transtegumental absorption of a neutral amino acid. It is concluded from these experiments that Diclidophora has a tegumental transport system for absorbing certain neutral amino acids, and whilst, clearly, the worm is sanguinivorous and digests blood in a well-developed gut, it may also be capable of supplementing this diet with low molecular weight organic nutrient absorbed directly from sea water via the tegument.

scholarly journals The sites of hydrolysis of dipeptides containing leucine and glycine by rat jejunum in vitro

1969 ◽  
Vol 114 (4) ◽  
pp. 855-861 ◽  
Author(s):  
E. B. Fern ◽  
R. C. Hider ◽  
D. R. London
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
The Other ◽  
Peptidase Activity ◽  
Rat Jejunum ◽  
Effective Inhibitor ◽  
Inhibitory Effects ◽  
Molar Basis ◽  
Hydrolysis Of

1. The effect of peptides containing leucine and glycine on accumulation of leucine and glycine by everted jejunal rings was studied. 2. It was shown that, on a molar basis, leucyl-leucine is a more effective inhibitor of uptake of [14C]leucine than is either leucylglycine or glycyl-leucine. These latter dipeptides behave alike. 3. The concentration of the dipeptides and their constituent amino acids in both the incubation medium and the tissue has been followed in these experiments by amino acid analysis. No leucine-containing peptides were observed in the tissue. 4. The inhibitory effects of the mixed dipeptides are altered by pH changes in an analogous way to the alterations in peptidase activity. 5. The experimental results indicate that leucine-containing peptides are hydrolysed before the transport step. 6. Glycylglycine, on the other hand, has only a small effect on the accumulation of glycine, although large amounts of the peptide accumulate unchanged in the tissue. This suggests that glycylglycine is taken up by a different mechanism to that for the leucine dipeptides.

scholarly journals ywfE in Bacillus subtilis Codes for a Novel Enzyme, l-Amino Acid Ligase

2005 ◽  
Vol 187 (15) ◽  
pp. 5195-5202 ◽  
Author(s):  
Kazuhiko Tabata ◽  
Hajime Ikeda ◽  
Shin-ichi Hashimoto
Keyword(s):  
Amino Acids ◽  
Bacillus Subtilis ◽  
Amino Acid ◽  
High Performance ◽  
Consensus Sequence ◽  
Reaction Products ◽  
In Silico Screening ◽  
Vitro Assay ◽  
Hydrolysis Of

ABSTRACT The ATP-dependent carboxylate-amine/thiol ligase superfamily is known to contain enzymes catalyzing the formation of various types of peptide, such as d-alanyl-d-alanine, polyglutamate, and γ-peptide, but, curiously, no enzyme synthesizing α-dipeptides of l-amino acids is known. We attempted to find such an enzyme. By in silico screening based on the consensus sequence of the superfamily followed by an in vitro assay with purified enzyme to avoid the degradation of the peptide(s) synthesized, ywfE of Bacillus subtilis was found to code for the activity forming l-alanyl-l-glutamine from l-alanine and l-glutamine with hydrolysis of ATP to ADP. No AMP was formed, supporting the idea that the enzyme belongs to the superfamily. Surprisingly, the enzyme accepted a wide variety of l-amino acids. Among 231 combinations of l-amino acids tested, reaction products were obtained for 111 combinations and 44 kinds of α-dipeptides were confirmed by high-performance liquid chromatography analyses, while no tripeptide or longer peptide was detected and the d-amino acids were inert. From these results, we propose that ywfE encodes a new member of the superfamily, l-amino acid ligase.

scholarly journals Effects of p-chlorophenylalanine and α-methylphenylalanine on amino acid uptake and protein synthesis in mouse neuroblastoma cells

1978 ◽  
Vol 174 (3) ◽  
pp. 931-938 ◽  
Author(s):  
C J Kelly ◽  
T C Johnson
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Animal Models ◽  
Phenylalanine Hydroxylase ◽  
Present Report ◽  
Neuroblastoma Cells ◽  
Acid Uptake

The phenylalanine analogues p-chlorophenylalanine and alpha-methylphenylalanine were used to inhibit phenylalanine hydroxylase in animal models for phenylketonuria. The present report examines the affects of these analogues on the metabolism of neuroblastoma cells. p-Chlorophenylalanine inhibited growth and was toxic to neuroblastoma cells. Although in vivo this analogue increased cell monoribosomes by 42%, it did not significantly affect poly(U)-directed protein synthesis in vitro. P-Chlorophenylalanine did not compete with phenylalanine or tyrosine for aminoacylation of tRNA and was therefore not substituted for those amino acids in nascent polypeptides. The initial cellular uptake of various large neutral amino acids was inhibited by this analogue but did not affect the flux of amino acids already in the cell; this suggested that an alteration of the cell's amino acid pools was not responsible for the cytotoxicity of the analogues. In contrast with p-chlorophenylalanine, alpha-methylphenylalanine did not exert these direct toxic effects because the administration of alpha-methylphenylalanine in vivo did not affect brain polyribosomes and a comparable concentration of this analogue was neither growth inhibitory nor cytotoxic to neuroblastoma cells in culture. The suitability of each analogue as an inhibitor of phenylalanine hydroxylase in animal models for phenylketonuria is discussed.

scholarly journals SUN-266 Protein Induced Pancreatic Hormone Secretion Is Modulated by Vagal CaSR

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Mariana Norton ◽  
Simon C Cork ◽  
Aldara Martin Alonso ◽  
Anna G Roberts ◽  
Yateen S Patel ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Hormone Secretion ◽  
Sensory Information ◽  
Glucagon Secretion ◽  
Vagal Afferents ◽  
Acid Uptake

Abstract The existence of a vago-vagal entero-pancreatic pathway, where sensory information from the gut can signal via vagal afferents to the brain to mediate changes in pancreatic function, has been recognised for over a century, and investigated extensively with regards to pancreatic exocrine secretions. However, the role of such pathways in pancreatic endocrine secretions has received less attention. The secretion of insulin and glucagon in response to protein and amino acids is conserved across species. This effect is thought to promote amino acid uptake into tissues without concomitant hypoglycaemia. We found that the essential amino acid L-Phenylalanine potently stimulates glucagon secretion, even when administered directly into the gut at small doses unlikely to significantly raise systematic levels. Administration of L-Phenylalanine also increased neuronal activation in the rat and mouse dorsal vagal complex, the central nervous system region directly innervated by vagal afferents. L-Phenylalanine modulates the activity of the calcium sensing receptor (CaSR), a nutrient sensor more commonly known for its role in calcium homeostasis, but which is thought to also act as a sensor of aromatic amino acids. Interestingly, the CaSR is one of the few nutrient sensors expressed in vagal afferents and in vitro calcium imaging revealed CaSR synthetic agonists activate subpopulations of vagal afferents. The role of CaSR in vivo was investigated further by selectively knocking down the CaSR in vagal afferents. Briefly, CaSR floxed mice were bilaterally injected directly into the nodose ganglion, where the cell bodies of vagal afferents are located, with a cre expressing adeno-associated virus. CaSR knockdown did not interfere with normal food intake, nor the vagal-dependent anorectic effects of cholecystokinin, or of L-Phenylalanine. However, it did blunt protein-induced glucagon secretion, suggesting involvement of the CaSR in the vagus nerve in protein sensing and glucose homeostasis. Future studies are required to determine the importance of vagal CaSR in protein induced pancreatic endocrine secretions, and the possibility of exploiting this circuit to develop new anti-diabetic therapies.

scholarly journals Bacillus coagulans GBI-30, 6086 improves amino acid absorption from milk protein

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Richard A. Stecker ◽  
Jessica M. Moon ◽  
Travis J. Russo ◽  
Kayla M. Ratliff ◽  
Petey W. Mumford ◽  
...  
Keyword(s):  
Amino Acids ◽  
Small Intestine ◽  
Amino Acid ◽  
Milk Protein ◽  
Bacillus Coagulans ◽  
Protein Concentrate ◽  
Protein Digestion ◽  
Milk Protein Concentrate ◽  
Mean Differences

Abstract Background Probiotic Bacillus coagulans GBI-30, 6086 (BC30) has been shown to increase protein digestion in an in vitro model of the stomach and small intestine. Once active in the small intestine after germination, BC30 aids the digestion of carbohydrates and proteins. The extent to which BC30 administration may impact protein digestion and amino acid appearance in humans after protein ingestion is currently unknown. This study examined the impact of adding BC30 to a 25-g dose of milk protein concentrate on post-prandial changes in blood amino acids concentrations. Methods 14 males and 16 females (n = 30, 26.4 ± 6.5 years; 172.3 ± 10.8 cm; 78.2 ± 14.8 kg; 22.6 ± 7.2% fat) completed two supplementation protocols that each spanned two weeks separated by a washout period that lasted three weeks. Participants were instructed to track their dietary intake and ingest a daily 25-g dose of milk protein concentrate with (MPCBC30) or without (MPC) the addition of BC30. Body composition and demographics were assessed upon arrival to the laboratory. Upon ingestion of their final assigned supplemental dose, blood samples were taken at 0 (baseline), 30, 60, 90, 120, 180, and 240 min post-consumption and analyzed for amino acid concentrations. Results Arginine (p = 0.03) and Isoleucine (p = 0.05) revealed greater area-under-the curve (AUC) in MPCBC30 group compared to MPC. In addition, Arginine (p = 0.02), Serine (p = 0.01), Ornithine (p = 0.02), Methionine (p = 0.04), Glutamic Acid (p = 0.01), Phenylalanine (p = 0.05), Isoleucine (p = 0.04), Tyrosine (p = 0.02), Essential Amino Acids (p = 0.02), and Total Amino Acids (p < 0.01) all revealed significantly greater concentration maximum (CMax) in MPCBC30 compared to MPC. Finally, time to reach CMax (TMax) was significantly faster for Glutamine (p < 0.01), Citrulline (p < 0.01), Threonine (p = 0.04), Alanine (p = 0.02) in MPCBC30 when compared to MPC. Greater mean differences between groups for AUC and CMax in women when compared to the mean differences in men were found for several amino acids. Conclusion In concert with previous in vitro evidence of improved protein digestion and amino acid appearance, these results reveal that adding BC30 to protein sources such as milk protein concentrate can improve AUC, CMax, and faster TMax. Follow-up research should examine differences between gender and explore how aging can impact these outcomes. Retrospectively registered on June 11, 2020 at ClinicalTrials.gov as NCT04427020.

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