The influence of bovine colostrum supplementation on exercise performance in highly trained cyclists

Purpose: The aim of this experiment was to investigate the influence of low dose bovine colostrum supplementation on exercise performance in cyclists over a 10 week period that included 5 days of high intensity training (HIT). Methods: Over 7 days of preliminary testing, 29 highly trained male road cyclists completed a VO2max test (in which their ventilatory threshold was estimated), a time to fatigue test at 110% of ventilatory threshold, and a 40 km time trial (TT40). Cyclists were then assigned to either a supplement (n = 14, 10 g/day bovine colostrum protein concentrate (CPC)) or a placebo group (n = 15, 10 g/day whey protein) and resumed their normal training. Following 5 weeks of supplementation, the cyclists returned to the laboratory to complete a second series of performance testing (week 7). They then underwent five consecutive days of HIT (week 8) followed by a further series of performance tests (week 9). Results: The influence of bovine CPC on TT40 performance during normal training was unclear (week 7: 1+/-3.1%, week 9: 0.1+/-2.1%; mean+/-90% confidence limits). However, at the end of the HIT period, bovine CPC supplementation, compared to the placebo, elicited a 1.9+/-2.2% improvement from baseline in TT40 performance and a 2.3+/-6.0% increase in time trial intensity (% VO2max), and maintained TT40 heart rate (2.5+/-3.7%). In addition, bovine CPC supplementation prevented a decrease in ventilatory threshold following the HIT period (4.6+/-4.6%). Conclusion: Low dose bovine CPC supplementation elicited improvements in TT40 performance during an HIT period and maintained ventilatory threshold following five consecutive days of HIT.

Crystal structure of mammalian purple acid phosphatase

Background: Mammalian purple acid phosphatases are highly conserved binuclear metal-containing enzymes produced by osteoclasts, the cells that resorb bone. The enzyme is a target for drug design because there is strong evidence that it is involved in bone resorption. Results: The 1.55 Angstrom resolution structure of pig purple acid phosphatase has been solved by multiple isomorphous replacement. The enzyme comprises two sandwiched beta sheets flanked by or-helical segments. The molecule shows internal symmetry, with the metal ions bound at the interface between the two halves. Conclusions: Despite less than 15% sequence identity, the protein fold resembles that of the catalytic domain of plant purple acid phosphatase and some serine/threonine protein phosphatases. The active-site regions of the mammalian and plant purple acid phosphatases differ significantly, however. The internal symmetry suggests that the binuclear centre evolved as a result of the combination of mononuclear ancestors. The structure of the mammalian enzyme provides a basis for antiosteoporotic drug design.

Tetrahydrofolates are greatly stabilized by binding to bovine milk folate-binding protein

The dietary supply of folates and their measurement are both affected, potentially, by the instability of some folates. Labile folates appear to be stabilized by binding to folate-binding protein (FBP); this paper reports measurements of that stabilization. The degradation rates of the very labile tetrahydrofolate (H(4)folate) and moderately labile 5-methyltetrahydrofolate (5-CH(3)H(4)folate) were measured with the compounds free or bound to either soluble or immobilized bovine milk FBP. Complexation increased stability from 2- to > 1000-fold, depending on buffer and temperature conditions. H(4)folate at 4degreesC and pH 6.7 appeared to be quite stable for > 100 d when bound to soluble FBP but had a half-life of < 1 h when free. Stabilization of milk folates may be a role of FBP and would improve the bioavailability of milk folate to newborns and other consumers.

The effects of level of dietary protein on the milk production and rumen physiology of dairy cows fed a diet based on a tropical grass hay

zFour rumen-fistulated, multiparous Holstein-Friesian cows in early lactation were offered mixed diets based on rhodes grass hay (Chloris gayana) cv. Callide containing 13, 14, 15 or 16% crude protein (CP) on a dry matter basis, in a 4 x 4 latin square design. The estimated undegradable protein concentration in these diets was similar with rumen degradable protein concentration varying. Cows fed a diet containing 13% CP had lower (P = 0.07) milk yields than cows in other treatments (20.4 vs 21.9, 22.0 and 22.2 L/d for 13, 14, 15 and 16% CP, respectively). A positive linear relationship was found (P = 0.06) between organic matter intake and dietary CP%. There were negative linear relationships between dietary CP% and digestibilities of dry matter (P = 0.09), organic matter (P = 0.06) and neutral detergent fibre (P = 0.02). Feeding a diet containing 13% CP resulted in significantly lower (P = 0.001) molar proportions (%) of rumen valerate in comparison with other treatments. The molar proportions of isovalerate differed (P = 0.001) between treatments (0.66, 0.78, 0.89 and 1.04%) for 13, 14, 15 and 16% CP, respectively). Dietary protein level had no effect on rates of passage, in situ digestion of rhodes grass hay or ratios of allantoin: creatinine in urine. These data showed that increasing the dietary CP concentration of lactating cows fed diets based on rhodes grass hay increased intakes and not significantly improved at dietary CP concentrations above 14% DM.

Dietary interactions influence the effects of bovine folate-binding protein on the bioavailability of tetrahydrofolates in rats

The newborns of mammals have a high folate demand, yet obtain adequate folate nutrition solely from their mothers' milk despite its low folate content. Milk folate is entirely bound by an excess of folate-binding protein (FBP), prompting speculation that FBP may affect the bioavailability of the limited folate supply. Previous research has shown that FBP-bound folic acid is more gradually absorbed, thereby reducing the peak plasma folate concentration and preventing loss into the urine. Natural folates are reduced derivatives of folic acid, with milk predominantly containing 5-methyltetrahydrofolate, yet little research has been carried out to determine the role of FBP in the bioavailability of reduced folates. We studied the effect of FBP on folate nutrition of rats in both single-dose and 4-wk feeding experiments. The effect of FBP was influenced by the presence of other milk components. FBP increased bioavailability of dietary folate when it was consumed with other whey proteins or with soluble casein. However, in the presence of acid-precipitated casein or a whey preparation enriched in lipids, bioavailability was decreased. These results highlight the difficulties of extrapolating from experimental results obtained using purified diets alone and of studying interactions among dietary components. They suggest that the addition of FBP-rich foods to folate-rich foods could enhance the bioavailability of natural folates, but that the outcome of such a combination would depend on interactions with other components of the diet.

The effect of different levels of dietary crude protein on urea metabolism of rusa deer (Cervus timorensis)

Two experiments were conducted to measure urea recycling and rumen flow dynamics in young rusa deer fed low (LP) or high (HP) protein diets. Pool size and flux rate of labelled urea. into and out of the blood pool were measured using single intravenous (i.v.) injection solutions containing [C-14] - and [N-15]-urea. A curve peeling technique was used to fit the enrichment of N-15 or specific radioactivity (SRA) of C-14 to exponential equations. Body urea-N pool size was significantly greater (P < 0.05) when a HP, compared to a LP diet, was fed. Urea space, expressed as a percent of live weight, total flux rate of urea through the blood pool and the irreversible loss of urea was similar for both diets. The mean (+/- S.E.M.) concentration of plasma urea-N was greater when animals were fed the HP diet compared to the LP diet (2 1.1 +/- 0.3 versus 14.4 +/- 1.4 mg/100 ml, respectively). Voluntary feed intake and digestibility of dietary components were also measured. Daily dry matter intakes were not affected by the crude protein (CP) content of the diet, although apparent DM digestibility was significantly greater for HP diet fed in both experiments. An intraruminal infusion of CrEDTA was used to determine rumen flow dynamics. Ruminal mean retention time, relative net outflow rate of water and passage rate constant (k(w)) were significantly greater (P < 0.05) when the HP diet was fed compared to the LP diet. The extent of urea metabolism and flux rates of urea between the blood and secondary pools appear similar to those previously reported for other ruminants fed diets contrasting in CP content. (c) 2005 Elsevier B.V. All rights reserved.

Low dietary protein during early pregnancy alters bovine placental development

To determine if low dietary protein concentration in the first two trimesters of pregnancy alters placental development, genetically similar heifers from closed herd were fed diets containing different levels of protein in the first and second trimesters of gestation. There were four animals per treatment group, the groups being: L/L = fed a diet containing 7% crude protein (CP) (low protein) in the first and second trimesters; H/H = fed a diet containing 14% (P thigh protein) in the first and second trimesters; L/H = fed low protein in the first trimester and high in the second trimester and vice versa for the H/L group. Low protein diets in the first trimester increased dry cotyledon weight at term. Trophectoderm volume density increased in the H/L and L/H group compared to the L/L and H/H groups. Blood vessel volume and volume density in foetal villi decreased in the H/L and L/H groups compared with the H/H and L/L groups. There was no effect of diet treatment on cotyledon number, diameter or wet weight and no effect on the volume density of connective tissue or fibroblasts in the foetal villi. These results show that a low dietary protein concentration in the first trimester of pregnancy followed by increased protein in the second trimester enhanced placental development. Further, trophectoderm volume was highly correlated with birth weight. Early protein restriction in the pregnant cow may enhance foetal growth in part by stimulating placental growth and function. (C) 1999 Published by Elsevier Science B.V. All rights reserved.

DNA packaging by L1 and L2 capsid proteins of bovine papillomavirus type 1

Encapsidation of circular DNA by papillomavirus capsid protein was investigated in Cos-1 cells. Plasmids carrying both an SV40 origin of replication (or) and an E. coli on were introduced into Cos-1 cells by DNA transfection. PV capsid proteins were supplied in trans by recombinant vaccinia viruses. Pseudovirions were purified from infected cells and their packaged DNA was extracted and used to transform E. coil as an indication of packaging efficacy. VLPs assembled from BPV-1 L1 alone packaged little plasmid DNA, whereas VLPs assembled from BPV-1 L1+L2 packaged plasmid DNA at least 50 times more effectively. BPV-1 L1+L2 VLPs packaged a plasmid containing BPV-1 sequence 8.2 +/- 3.1 times more effectively than a plasmid without BW sequences. Using a series of plasmid constructs comprising a core BPV-1 sequence and spacer DNA it was demonstrated that BW VLPs could accommodate a maximum of about 10.2 kb of plasmid DNA, and that longer closed circular DNA was truncated to produce less dense virions with shorter plasmid sequences. The present study suggests that packaging of genome within PV virions involves interaction of L2 protein with specific DNA sequences, and demonstrates that PV pseudovirions have the potential to be used as DNA delivery vectors for plasmids of up to 10.2 kb. (C) 1998 Academic Press.

Purification,characterization and crystallization in two crystal forms of bovine cyclophilin 40

The purification and crystallization of two different crystal forms of the two-domain protein bovine cyclophilin 40 is reported. Tetragonal crystals grown in methyl pentanediol belong to space group P4(2)22 with unit-cell parameters a = 94.5, c = 118.3 Angstrom. Long thin needles grown from PEG belong to space group C2 with unit-cell parameters a = 125.71, b = 47.3, c = 74.6 Angstrom, beta = 93.90 degrees. The N-terminal 170 amino acids have significant homology with the well characterized human cyclophilin A. The C-terminal domain is largely made up of three copies of the tetratricopeptide repeat motif thought to be involved in mediating protein-protein interactions. Cyclophilins are frequently found as domains in larger multidomain proteins. To date, only X-ray structures of single-domain cyclophilins have been reported, and this work provides the first example of the purification and crystallization of a larger protein containing a cyclophilin domain.

Post translational modifications of recombinant human Papillomavirus type 6b major capsid protein

We have determined the post-translational modifications of the major capsid protein, L1 of human papillomavirus (HPV) type 6b. Since this virus cannot be cultured in the laboratory to obtain sufficient material for a study, a recombinant L1 protein produced in a vaccinia virus expression system was used in this investigation. Our results show that this protein is phosphorylated at serine residues and is also glycosylated. No myristoylation or palmitoylation was detected. The fraction of L1 protein incorporated into virus-like particles was not glycosylated. Since recombinant L1 protein is a potential human vaccine candidate, knowledge of the post-translation modifications of this protein may prove useful for the design of anti-HPV vaccines. (C) 1999 Elsevier Science B.V. All rights reserved.

Active site-directed protein regulation

Regulation of protein function is vital for the control of cellular processes. Proteins are often regulated by allosteric mechanisms, in which effecters bind to regulatory sites distinct from the active sites and alter protein function. Intrasteric regulation, directed at the active site and thus the counterpart of allosteric control, is now emerging as an important regulatory mechanism.

Differences in the post-translational modifications of human papillomavirus type 6b major capsid protein expressed from a baculovirus system compared with a vaccinia virus system

Virus-like particles (VLPs) are being currently investigated in vaccines against viral infections in humans. There are different recombinant-protein-expression systems available for obtaining the necessary VLP preparation for vaccination. However, the differences in post-translational modifications of the recombinant proteins obtained and their differences in efficacy in eliciting an anti-viral response in vaccines are not well established. In this study we have compared the posttranslational modifications of human papillomavirus type-6b major capsid protein L1 (HPV 6bL1) expressed using recombinant baculovirus (rBV) in Sf9 (Spodoptera frugiperda) insect cells, with the protein expressed using recombinant vaccinia virus (rVV) in CV-1 kidney epithelial cells, Two-dimensional gel electrophoresis of biosynthetically labelled rBV-expressed HPV 6bL1 showed several post-translationally modified variants of the protein, whereas rVV-expressed HPV 6bL1 showed only a few variants. Phosphorylations were detected at threonine and serine residues for the L1 expressed from rBV compared with phosphorylation at serine residues only for the L1 expressed from rVV. HPV 6bL1 expressed using rBV incorporated [H-3]mannose and [H-3]galactose, whereas HPV 6bL1 expressed using rVV incorporated only [H-3]galactose. We conclude that post-translational modification of recombinant HPV 6bL1 can differ according to the system used for its expression. Since recombinant L1 protein is a potential human-vaccine candidate, the implication of the observed differences in post-translational modifications on immunogenicity of L1 VLPs warrants investigation.