scholarly journals Dietary Phenylalanine and Tyrosine Requirements in Healthy Human Pregnancy

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 978-978
Author(s):  
Madeleine Ennis ◽  
Kenneth Lim ◽  
Ronald Ball ◽  
Paul Pencharz ◽  
Glenda Courtney-Martin ◽  
...  
Keyword(s):  
Amino Acid ◽  
Life Stage ◽  
Experimental Studies ◽  
Late Pregnancy ◽  
Acid Oxidation ◽  
Current Recommendation ◽  
Two Phase ◽  
Healthy Human ◽  
Dietary Requirement ◽  
Indispensable Amino Acid

Abstract Objectives Phenylalanine (PHE), an indispensable amino acid, is required for protein synthesis and is fundamental for proper fetal development. PHE is the precursor for tyrosine (TYR), a conditionally indispensable amino acid that is crucial for synthesis of dopamine, norepinephrine and epinephrine. The current recommendation for PHE + TYR during pregnancy is 36 mg/kg/d, which is factorially determined and not based on experimental studies. Our objective was to experimentally determine PHE + TYR requirements during early (13–19 wks) and late (33–39 wks) gestation, accounting for potential changes in requirement since metabolism and rate of growth are not static throughout pregnancy. Methods 20 women (22–39 y) were studied using the non-invasive stable isotope based indicator amino acid oxidation technique (with L-[1–,13C]Leucine as indicator) for a total of 51 study days. A range of PHE intakes (5 to 100 mg/kg/d) were provided in early and late pregnancy as isonitrogenous and isocaloric meals on separate study days, in the absence of dietary tyrosine. Breath samples were collected at baseline and isotopic steady state. Samples were analyzed for ,13C enrichment on an Isotope Ratio Mass Spectrometer, and the PHE + TYR requirement was determined using two-phase linear regression crossover analysis to identify a breakpoint in ,13CO2 production (represents the minimum dietary requirement) in response to changes in PHE intake. Results Results suggest the PHE + TYR requirement is 43 and 55 mg/kg/d in early and late pregnancy respectively. Conclusions Our preliminary results suggest that PHE + TYR requirements in pregnancy are higher compared to current recommendations, and are different between gestational stages. These results are similar to our previous findings of increased requirements for dietary protein, lysine and PHE (in the presence of excess TYR) in late compared to early pregnancy. They also highlight the need to investigate the dietary requirement of the remaining indispensable amino acids during this life stage. Funding Sources Canadian Institute of Health Research.

scholarly journals Phenylalanine requirement in children with classical PKU determined by indicator amino acid oxidation

2002 ◽  
Vol 283 (6) ◽  
pp. E1249-E1256 ◽  
Author(s):  
Glenda Courtney-Martin ◽  
Rachelle Bross ◽  
Mahroukh Raffi ◽  
Joe T. R. Clarke ◽  
Ronald O. Ball ◽  
...  
Keyword(s):  
Amino Acid ◽  
Acid Oxidation ◽  
Phenylalanine Concentration ◽  
Two Phase ◽  
Amino Acid Oxidation ◽  
Blood Phenylalanine ◽  
Plasma Phenylalanine ◽  
Crossover Analysis ◽  
The Mean ◽  
The Difference

Dietary restriction of phenylalanine is the main treatment for phenylketonuria (PKU), and current estimates of requirements are based on plasma phenylalanine concentration and growth. The present study aimed to determine more precisely the phenylalanine requirements in patients with the disease by use of indicator amino acid oxidation, withl-[1-13C]lysine as the indicator. Breath13CO2 production (F13 co 2) was used as the end point. Finger-prick blood samples were also collected for measurement of phenylalanine to relate phenylalanine intake to blood phenylalanine levels. The mean phenylalanine requirement, estimated using a two-phase linear regression crossover analysis, was 14 mg · kg−1 · day−1, and the safe population intake (upper 95% confidence interval of the mean) was found to be 19.5 mg · kg−1 · day−1. A balance between phenylalanine intake and the difference between fed and fasted blood phenylalanine concentration was observed at an intake of 20 mg · kg−1 · day−1. The similarity between these two values (19.5 and 20 mg · kg−1 · day−1) suggests that the maximal phenylalanine intake for children with PKU should be no higher than 20 mg · kg−1 · day−1.

scholarly journals Adult dogs of different breed sizes have similar threonine requirements as determined by the indicator amino acid oxidation technique

2020 ◽  
Vol 98 (3) ◽  
Author(s):  
Wilfredo D Mansilla ◽  
Lisa Fortener ◽  
James R Templeman ◽  
Anna K Shoveller
Keyword(s):  
Amino Acid ◽  
Basal Diet ◽  
Acid Oxidation ◽  
Total Production ◽  
Experimental Diet ◽  
Two Phase ◽  
Deficient Diet ◽  
Amino Acid Oxidation ◽  
The Mean ◽  
Labrador Retrievers

Abstract Threonine (Thr) requirements for immature (growing) Beagles have been determined, but little knowledge is available on Thr requirements for maintenance in mature dogs. Moreover, differences of Thr requirements among different breeds or sizes of adult dogs have not been investigated. The objective of the present study was to determine Thr requirements in adult dogs of three different breeds using the indicator amino acid oxidation (IAAO) technique. In total, 13 adult dogs were used, 4 Miniature Dachshunds (5.8 ± 0.4 kg body weight [BW]; 3 spayed and 1 neutered), 4 spayed Beagles (9.3 ± 0.6 kg BW), and 5 neutered Labrador Retrievers (30.5 ± 1.7 kg BW). Dogs were fed a Thr-deficient diet (Thr = 0.23%) and randomly allocated to receiving one of seven concentrations of Thr supplementation (final Thr concentration in experimental diets was 0.23%, 0.33%, 0.43%, 0.53%, 0.63%, 0.73%, and 0.83%; as fed basis) for 2 d. After 2 d of adaptation to the experimental diets, dogs underwent individual IAAO studies. During the IAAO studies, total daily feed was divided into 13 equal meals; at the sixth meal, dogs were fed a bolus of l-[1-13C]-Phenylalanine (Phe) (9.40 mg/kg BW), and thereafter, l-[1-13C]-Phe (2.4 mg/kg BW) was supplied with every meal. Before feeding the next experimental diet, dogs were fed a Thr-adequate basal diet for 4 d (Thr = 0.80% as fed basis) in known amounts that maintained individual dog BW. Total production of 13CO2 during isotopic steady state was determined by enrichment of 13CO2 in breath samples and total production of CO2 measured using indirect calorimetry. The mean requirements for Thr, defined as the breakpoint, and the 95% confidence interval (CI) were determined using a two-phase linear regression model. For Miniature Dachshunds, the two-phase model was not significant, and Thr requirements could not be determined. Mean Thr requirements for Beagles and Labradors were 72.2 and 64.1 mg/kg BW on an as-fed basis, respectively. The requirement for Thr between these two dog breeds was not different (P > 0.10). Thus, the data for Beagles and Labradors were pooled and a mean requirement for Thr was determined at 66.9 mg/kg BW, and the 95% CI was estimated at 84.3 mg/kg BW. In conclusion, estimated Thr requirements for Beagles and Labradors did not differ, and these recommendations are higher than those suggested by NRC (2006) and AAFCO (2014) for adult dogs at maintenance.

scholarly journals Lysine requirements in small, medium, and large breed adult dogs using the indicator amino acid oxidation technique1

2020 ◽  
Vol 4 (3) ◽  
Author(s):  
Katja A K Sutherland ◽  
Wilfredo D Mansilla ◽  
Lisa Fortener ◽  
Anna K Shoveller
Keyword(s):  
Amino Acid ◽  
Feed Intake ◽  
Dry Matter ◽  
National Research Council ◽  
Acid Oxidation ◽  
Total Production ◽  
Test Diet ◽  
Two Phase ◽  
Amino Acid Oxidation ◽  
Matter Basis

Abstract There is a lack of knowledge regarding the lysine (Lys) requirements of mature dogs and whether there are breed differences. The present study aimed to determine the Lys requirement in three breeds of mature dogs using the indicator amino acid oxidation (IAAO) technique. Thirteen adult dogs were used, four Miniature Dachshunds (5.39 ± 0.71 kg; 1.05 ± 0.02 yr old, mean ± SD), four Beagles (8.09 ± 0.40 kg; 5.03 ± 0.09 yr old, mean ± SD), and five Labrador Retrievers (29.42 ± 2.04 kg; 3.30 ± 0.69 yr old, mean ± SD). After 14 d of adaptation to a basal extruded kibble diet, dogs were fed a test diet mildly deficient in Lys (Lys concentration = 0.36%) at 17 (Miniature Dachshunds) or 13 g/kg body weight (BW; Beagles and Labradors) for 2 d. The test diet was supplemented with one of seven isonitrogenous Lys–Ala solutions, resulting in a final dietary Lys concentration of 0.36%, 0.40%, 0.44%, 0.50%, 0.54%, 0.58%, and 0.62% (as-fed basis). Dogs received dietary concentrations of Lys in random order and no dog received the same order. Following 2 d of adaptation to the experimental diets, the dogs underwent IAAO studies. During the IAAO studies, total daily feed was divided in 13 equal meals. At the sixth meal, dogs were fed a bolus of L-[1-13C]-Phe (9.40 mg/kg BW); thereafter, L-[1-13C]-Phe was supplied with every meal (2.4 mg/kg BW). Total production of 13CO2 (F13CO2) during isotopic steady state was determined by enrichment of 13CO2 of breath samples and total production of CO2, measured using indirect calorimetry. A two-phase linear regression model was used to derive the mean Lys requirement, defined as the breakpoint, and the upper 95% confidence limit was calculated as the recommended allowance (RA) for Lys intake. For Miniature Dachshunds, the study was repeated with a feed intake of 14 g/kg BW, but Lys requirements could not be determined at either feed intake, suggesting a requirement below the lowest concentration and intake. Mean Lys requirements for Beagles and Labradors were 0.455% (59.16 mg/kg BW) and 0.440% (57.19 mg/kg BW), respectively, on a dry matter basis. Pooling the data for these breeds provides a mean estimate of the Lys requirement at 0.448% (58.21 mg/kg BW) with an upper 95% CL of 0.526% (68.41 mg/kg BW) on a dry matter basis. In conclusion, the Lys requirements of Beagles and Labradors are similar, while the requirement for Miniature Dachshunds is undetermined and likely lower. The estimated Lys requirement for Beagles and Labradors is higher than the National Research Council recommendation.

scholarly journals Tryptophan requirements in small, medium, and large breed adult dogs using the indicator amino acid oxidation technique1

2019 ◽  
Vol 97 (8) ◽  
pp. 3274-3285 ◽  
Author(s):  
James R Templeman ◽  
Wilfredo D Mansilla ◽  
Lisa Fortener ◽  
Anna K Shoveller
Keyword(s):  
Amino Acid ◽  
Dry Matter ◽  
Basal Diet ◽  
Acid Oxidation ◽  
Total Production ◽  
Deficient Diet ◽  
Study Objective ◽  
Amino Acid Oxidation ◽  
Indispensable Amino Acid ◽  
Labrador Retrievers

Abstract Tryptophan (Trp) is an indispensable amino acid (AA) for dogs of all life stages; however, although Trp requirements for growing dogs are derived from 3 dose–response studies, there are no empirical data on Trp requirements for adult dogs at maintenance. The study objective was to determine Trp requirements of adult dogs of 3 different breeds using the indicator amino acid oxidation (IAAO) technique. Four spayed or neutered Miniature Dachshunds (5.28 ± 0.29 kg BW), 4 spayed Beagles (9.32 ± 0.41 kg BW), and 5 neutered Labrador Retrievers (30.51 ± 2.09 kg BW) were used. After a 14-d adaptation to a Trp-adequate basal diet (Trp = 0.482% dry matter), all dogs were fed a mildly Trp-deficient diet for 2 d (Trp = 0.092% dry matter) before being randomly allocated to receiving 1 of 7 concentrations of Trp supplementation (final Trp content in experimental diets was 0.092, 0.126, 0.148, 0.182, 0.216, 0.249, and 0.283% dry matter) and all dogs received all Trp treatments. After 2-d adaptation to the experimental diets, dogs underwent individual IAAO studies. Total feed was divided in 13 equal meals; at the sixth meal, dogs were fed a bolus of L-[1-13C]-Phenylalanine (Phe) (9.40 mg/kg BW), and thereafter, L-[1-13C]-Phe was supplied (2.4 mg/kg BW) with every meal. Total production of 13CO2 during isotopic steady state was determined by enrichment of 13CO2 in breath samples and total production of CO2 measured using indirect calorimetry. The maintenance requirement for Trp and the 95% confidence interval (CI) were determined using a 2-phase linear regression model. Mean Trp requirements were estimated at 0.154, 0.218, and 0.157% (dry-matter) for Dachshunds, Beagles, and Labradors, respectively. The upper 95% CI were 0.187, 0.269, and 0.204% (dry-matter) for Dachshunds, Beagles, and Labradors. In conclusion, estimated Trp requirements are higher for Beagles compared with Labradors or Dachshunds, and all estimated requirements are higher than those currently recommended by the NRC and AAFCO.

scholarly journals Development of a novel bioassay for determining the available lysine contents of foods and feedstuffs

2007 ◽  
Vol 20 (1) ◽  
pp. 3-16 ◽  
Author(s):  
Shane M. Rutherfurd ◽  
Paul J. Moughan
Keyword(s):  
Amino Acid ◽  
Nutritional Value ◽  
Acid Oxidation ◽  
Processed Foods ◽  
Lysine Content ◽  
Slope Ratio ◽  
Chemical Methods ◽  
Amino Acid Availability ◽  
Indispensable Amino Acid ◽  
Available Lysine

Lysine is an important indispensable amino acid, and describing the lysine content of a food or feedstuff provides useful information about nutritional value. However, when a food or feedstuff is subjected to heating the lysine present can be altered to nutritionally unavailable derivatives. These derivatives can revert back to lysine during the acid hydrolysis step used in amino acid analysis causing an overestimate of the lysine content. There have been many chemical methods developed to determine the reactive (unmodified) lysine content of foods and feedstuffs, but these do not take into account the incomplete absorption of lysine from the small intestine. There are also a number of animal-based assays for determining available lysine (the lysine that can be absorbed in a form that can be used for protein synthesis). The true ileal amino acid digestibility assay is commonly used to determine amino acid availability and is accurate for application to unprocessed foods and feedstuffs but is not accurate for lysine and possibly other amino acids when applied to heat-processed foods or feedstuffs. For such protein sources, assays such as the slope-ratio assay, indicator amino acid oxidation assay and the BIOLYSINE™ assay (true ileal digestible reactive lysine assay) have been developed to determine available lysine. The present paper discusses the efficacy of the BIOLYSINE™ assay as well as other assays for determining available lysine in processed foods and feedstuffs.

scholarly journals Dietary phenylalanine requirements during early and late gestation in healthy pregnant women

2019 ◽  
Vol 111 (2) ◽  
pp. 351-359 ◽  
Author(s):  
Madeleine A Ennis ◽  
Betina F Rasmussen ◽  
Kenneth Lim ◽  
Ronald O Ball ◽  
Paul B Pencharz ◽  
...  
Keyword(s):  
Amino Acid ◽  
Early Pregnancy ◽  
Resting Energy Expenditure ◽  
Late Pregnancy ◽  
Late Gestation ◽  
Acid Oxidation ◽  
Amino Acid Oxidation ◽  
Dietary Requirements ◽  
Isotope Ratio Mass Spectrometer ◽  
The Mean

ABSTRACT Background Phenylalanine is an indispensable amino acid and, via tyrosine, is the precursor for the neurotransmitters dopamine, norepinephrine, and epinephrine. Currently, dietary requirements for phenylalanine during pregnancy are unknown. Objectives This study's aim was to determine phenylalanine requirements (in the presence of excess tyrosine) during early and late gestation using direct amino acid oxidation (DAAO; with l-[1-13C]phenylalanine) and indicator amino acid oxidation (IAAO; with l-[1-13C]leucine). Methods Twenty-three healthy women (age: 30.4 ± 3.1 y, mean ± SD) were studied at a range of phenylalanine intakes (5.5–30.5 mg · kg−1 · d−1 in early and late pregnancy using DAAO, and 2.5–30.5 mg · kg−1 · d−1 in late pregnancy using IAAO) for a total of 76 study days. Test intakes were provided as 8 isocaloric and isonitrogenous meals with 1.5 g · kg−1 · d−1 protein and energy at 1.7 times the measured resting energy expenditure. Breath samples were analyzed on an isotope ratio mass spectrometer for 13C enrichment. Phenylalanine requirement was determined using a 2-phase linear regression crossover model to identify a breakpoint in 13CO2 production (representing the mean requirement) in response to phenylalanine intakes. Results Phenylalanine requirement during early pregnancy was determined to be 15 mg · kg−1 · d−1 (95% CI: 10.4, 19.9 mg · kg−1 · d−1); during late pregnancy, it was determined to be 21 mg · kg−1 · d−1 by DAAO (95% CI: 17.4, 24.7 mg · kg−1 · d−1) and IAAO (95% CI: 10.5, 32.2 mg · kg−1 · d−1). Conclusions Our results suggest a higher requirement (40%) for phenylalanine during late pregnancy than during early pregnancy. Moreover, the early pregnancy requirements are higher than the previous adult male requirement (9.1 mg · kg−1 · d−1; 95% CI: 4.6, 13.6 mg · kg−1 · d−1), although the 95% CIs overlap. Both DAAO and IAAO methods provided similar breakpoints in late pregnancy, showing that the DAAO method was appropriate even though low phenylalanine intakes could not be tested. These results have potential implications for gestation stage–specific dietary phenylalanine recommendations in future. This trial was registered at clinicaltrials.gov as NCT02669381.

scholarly journals Minimum dietary methionine requirements in Miniature Dachshund, Beagle, and Labrador Retriever adult dogs using the indicator amino acid oxidation technique

2020 ◽  
Vol 98 (11) ◽  
Author(s):  
Wilfredo D Mansilla ◽  
James R Templeman ◽  
Lisa Fortener ◽  
Anna K Shoveller
Keyword(s):  
Amino Acid ◽  
National Research Council ◽  
Basal Diet ◽  
Acid Oxidation ◽  
Total Production ◽  
Two Phase ◽  
Minimum Requirement ◽  
Linear Mixed Effects ◽  
Low Protein ◽  
Labrador Retrievers

Abstract The objective of this study was to determine the minimum requirement (MR) for methionine (Met), when cyst(e)ine (Cys) is provided in excess, in adult dogs of three different breed sizes using the indicator amino acid (AA) oxidation (IAAO) technique. In total, 12 adult dogs were used: 1 neutered and 3 spayed Miniature Dachshunds (4.8 ± 0.4 kg body weight [BW], mean ± SD), 4 spayed Beagles (9.5 ± 0.7 kg BW, mean ± SD), and 4 neutered Labrador Retrievers (31.8 ± 1.7 kg BW, mean ± SD). A deficient Met basal diet with excess Cys was formulated. Dogs were fed the basal diet randomly supplemented with different Met-Alanine (Ala) solutions to achieve final Met concentrations in experimental diets of 0.21%, 0.26%, 0.31%, 0.36%, 0.41%, 0.46%, and 0.66% (as-fed basis). After 2 d of adaptation to the experimental diets, dogs underwent individual IAAO studies. During the IAAO study day, the total feed was divided into 13 equal meals; at the sixth meal, dogs were fed a bolus of l-[1-13C]-phenylalanine (Phe), and thereafter, l-[1-13C]-Phe was supplied with every meal. The total production of 13CO2 during isotopic steady state was determined by the enrichment of 13CO2 in breath samples, and the total production of CO2 measured using indirect calorimetry. The mean MR for Met and the upper 95% confidence limit (CL) were determined using a two-phase linear mixed-effects regression model. For Miniature Dachshunds, the MR for Met was between the first two dietary Met concentrations and is, therefore, between 35.7 and 44.1 mg.kg BW−1·d−1 (0.21% to 0.26%, as-fed basis; no requirement could be determined on a metabolic BW basis). For Beagles and Labrador Retrievers, the MR for Met was 57.5 and 50.4 mg.kg BW−1·d−1, 107.7 and 121.8 mg/kg BW^0.75, or 0.338 and 0.360%, respectively (as-fed basis). The upper 95% CL of Met requirements was 77.9 and 72.4 mg.kg BW−1·d−1, 147.8 and 159.6 mg/kg BW^0.75,or 0.458 and 0.517% for Beagles, and Labradors, respectively (as-fed basis). When pooling data from Beagles and Labrador Retrievers, the MR and upper 95% CL were 56.0 and 75.8 mg.kg BW−1·d−1 or 118.4 and 150.5 mg/kg BW^0.75 or 0.360% and 0.482% (as-fed basis). In conclusion, the MR and the upper 95% CL for Met are different for Dachshunds when compared with Beagles and Labrador Retrievers. Using this low-protein diet, the estimated upper 95% CL Met requirement for Beagles and Labrador is higher than those recommended in the National Research Council (NRC), but NRC is similar to the estimated upper 95% CL for Dachshunds.

scholarly journals Dietary Aromatic Amino Acid Requirements During Early and Late Gestation in Healthy Pregnant Women

2020 ◽  
Vol 150 (12) ◽  
pp. 3224-3230
Author(s):  
Madeleine A Ennis ◽  
Anna-Joy Ong ◽  
Kenneth Lim ◽  
Ronald O Ball ◽  
Paul B Pencharz ◽  
...  
Keyword(s):  
Amino Acid ◽  
Pregnant Women ◽  
Early Pregnancy ◽  
Aromatic Amino Acids ◽  
Enrichment Analysis ◽  
Late Pregnancy ◽  
Late Gestation ◽  
Acid Oxidation ◽  
Plasma Phenylalanine ◽  
Amino Acid Requirements

ABSTRACT Background Phenylalanine and tyrosine (referred to as total aromatic amino acids; TAAs) are essential for protein synthesis, and are precursors for important catecholamines. Current estimated average requirement (EAR) recommendations for TAA during pregnancy are 36 mg·kg−1·d−1, and has not been experimentally determined. Objectives The aim was to determine TAA requirements (dietary phenylalanine in the absence of tyrosine) during early and late gestation using the indicator amino acid oxidation (IAAO, with L-[1-13C]leucine) technique. Methods Nineteen healthy pregnant women (age 22–38 y) were studied at a range of phenylalanine intakes (5 to 100 mg·kg−1·d−1) in early (13–19 wk) and/or late (33–39 wk) pregnancy for a total of 51 study days. Graded test intakes were provided as 8 hourly isonitrogenous and isocaloric meals. Breath samples were collected for 13C enrichment analysis on an isotope ratio mass spectrometer. A plasma sample was collected and analyzed for phenylalanine and tyrosine concentrations on an amino acid analyzer. The TAA requirement in early and late pregnancy was calculated using 2-phase linear regression crossover analysis that identified breakpoints in 13CO2 production (the requirement) in response to phenylalanine intakes. Results TAA requirement during early pregnancy was 44 mg·kg−1·d−1 (95% CI: 28.3, 58.8) and during late pregnancy was 50 mg·kg−1·d−1 (95% CI: 36.1, 63.1). In early and late pregnancy, plasma phenylalanine and tyrosine concentrations rose linearly in response to graded phenylalanine intakes. Conclusions Our results suggest that the current EAR of 36 mg·kg−1·d−1 for TAAs is underestimated. When compared with results previously determined in nonpregnant adults, early pregnancy requirements were similar (43 compared with 44 mg·kg−1·d−1, respectively). During late pregnancy, a 14% higher TAA requirement was observed when compared with early pregnancy. The results from this study have potential implications for creating gestation stage-specific TAA recommendations.

scholarly journals Phase of menstrual cycle affects lysine requirement in healthy women

2004 ◽  
Vol 287 (3) ◽  
pp. E489-E496 ◽  
Author(s):  
Wantanee Kriengsinyos ◽  
Linda J. Wykes ◽  
Laksiri A. Goonewardene ◽  
Ronald O. Ball ◽  
Paul B. Pencharz
Keyword(s):  
Amino Acid ◽  
Menstrual Cycle ◽  
Luteal Phase ◽  
Random Order ◽  
Follicular Phase ◽  
Acid Oxidation ◽  
Two Phase ◽  
Menstrual Cycles ◽  
17Β Estradiol ◽  
Healthy Females

The aim of this study was to investigate whether the phases of the menstrual cycle affect lysine requirement in healthy adult females, as determined by the indicator amino acid oxidation (IAAO) method. Five healthy females with regular menstrual cycles were studied at seven graded levels of lysine intake, in random order, with an oral [13C]phenylalanine tracer protocol in both the follicular and luteal phases. A total of 14 studies were conducted for each subject. Breath and plasma samples were collected according to the standard IAAO protocol. Serum 17β-estradiol and progesterone concentrations were measured on each IAAO study day. The rate of release of 13CO2 from [13C]phenylalanine oxidation (F13CO2) was measured, and a two-phase linear regression crossover model was applied to determine lysine requirement. F13CO2 was higher during the luteal phase ( P < 0.001) and was positively associated with serum concentrations of 17β-estradiol and progesterone. The F13CO2 data were adjusted for subjects and sex hormones and used to define breakpoints for lysine requirements. The lysine requirement of healthy females in the luteal phase was 37.7 mg·kg−1·day−1 and higher ( P = 0.025) than that of females in the follicular phase (35.0 mg·kg−1·day−1). At all lysine intake levels, plasma amino acids were lower and phenylalanine oxidation was higher in the luteal relative to the follicular phase. Therefore, we reason that the higher lysine requirement observed in the luteal phase is probably due to higher amino acid catabolism.

scholarly journals The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA

2021 ◽  
Vol 22 (12) ◽  
pp. 6490
Author(s):  
Olga A. Postnikova ◽  
Sheetal Uppal ◽  
Weiliang Huang ◽  
Maureen A. Kane ◽  
Rafael Villasmil ◽  
...  
Keyword(s):  
Amino Acid ◽  
Insertion Sequence ◽  
Experimental Studies ◽  
Spike Protein ◽  
Spike Glycoprotein ◽  
Furin Cleavage ◽  
New Host ◽  
S Protein ◽  
Furin Cleavage Site ◽  
Ribosome Pausing

The SARS-CoV-2 Spike glycoprotein (S protein) acquired a unique new 4 amino acid -PRRA- insertion sequence at amino acid residues (aa) 681–684 that forms a new furin cleavage site in S protein as well as several new glycosylation sites. We studied various statistical properties of the -PRRA- insertion at the RNA level (CCUCGGCGGGCA). The nucleotide composition and codon usage of this sequence are different from the rest of the SARS-CoV-2 genome. One of such features is two tandem CGG codons, although the CGG codon is the rarest codon in the SARS-CoV-2 genome. This suggests that the insertion sequence could cause ribosome pausing as the result of these rare codons. Due to population variants, the Nextstrain divergence measure of the CCU codon is extremely large. We cannot exclude that this divergence might affect host immune responses/effectiveness of SARS-CoV-2 vaccines, possibilities awaiting further investigation. Our experimental studies show that the expression level of original RNA sequence “wildtype” spike protein is much lower than for codon-optimized spike protein in all studied cell lines. Interestingly, the original spike sequence produces a higher titer of pseudoviral particles and a higher level of infection. Further mutagenesis experiments suggest that this dual-effect insert, comprised of a combination of overlapping translation pausing and furin sites, has allowed SARS-CoV-2 to infect its new host (human) more readily. This underlines the importance of ribosome pausing to allow efficient regulation of protein expression and also of cotranslational subdomain folding.

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