Amino acid analysis: Determination of cysteine plus half-cystine in proteins after hydrochloric acid hydrolysis with a disulfide compound as additive

Barkholt, Vibeke; Jensen, Arne L.

doi:10.1016/0003-2697(89)90059-6

Cited by 326 publications

(181 citation statements)

References 14 publications

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“…The fractions containing activity were analyzed by SDS-PAGE, pooled, concentrated (10-kDa Amicon filter; Millipore), and buffer exchanged to 20 mM 2-(N-morpholino)ethanesulfonic acid (MES)-NaOH, pH 6.5, 2 mM CaCl 2 , and 100 mM NaCl. The protein concentration was determined spectrophotometrically using the molar extinction coefficient ε 280 of 138,180 M Ϫ1 cm Ϫ1 as determined by amino acid analysis (4). The isoelectric point (pI) was determined by focusing on a PhastGel IEF 4 to 6.5 isoelectric focusing gel and a pI marker (2.8 to 6.5) (GE Healthcare) using the PhastSystem (Pharmacia, Uppsala, Sweden).…”

Section: Methodsmentioning

confidence: 99%

Enzymology and Structure of the GH13_31 Glucan 1,6-α-Glucosidase That Confers Isomaltooligosaccharide Utilization in the Probiotic Lactobacillus acidophilus NCFM

et al. 2012

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b Isomaltooligosaccharides (IMO) have been suggested as promising prebiotics that stimulate the growth of probiotic bacteria. Genomes of probiotic lactobacilli from the acidophilus group, as represented by Lactobacillus acidophilus NCFM, encode ␣-1,6 glucosidases of the family GH13_31 (glycoside hydrolase family 13 subfamily 31) that confer degradation of IMO. These genes reside frequently within maltooligosaccharide utilization operons, which include an ATP-binding cassette transporter and ␣-glucan active enzymes, e.g., maltogenic amylases and maltose phosphorylases, and they also occur separated from any carbohydrate transport or catabolism genes on the genomes of some acidophilus complex members, as in L. acidophilus NCFM. Besides the isolated locus encoding a GH13_31 enzyme, the ABC transporter and another GH13 in the maltooligosaccharide operon were induced in response to IMO or maltotetraose, as determined by reverse transcription-PCR (RT-PCR) transcriptional analysis, suggesting coregulation of ␣-1,6-and ␣-1,4-glucooligosaccharide utilization loci in L. acidophilus NCFM. The L. acidophilus NCFM GH13_31 (LaGH13_31) was produced recombinantly and shown to be a glucan 1,6-␣-glucosidase active on IMO and dextran and product-inhibited by glucose. The catalytic efficiency of LaGH13_31 on dextran and the dextran/panose (trisaccharide) efficiency ratio were the highest reported for this class of enzymes, suggesting higher affinity at distal substrate binding sites. The crystal structure of LaGH13_31 was determined to a resolution of 2.05 Å and revealed additional substrate contacts at the ؉2 subsite in LaGH13_31 compared to the GH13_31 from Streptococcus mutans (SmGH13_31), providing a possible structural rationale to the relatively high affinity for dextran. A comprehensive phylogenetic and activity motif analysis mapped IMO utilization enzymes from gut microbiota to rationalize preferential utilization of IMO by gut residents.

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Section: Methodsmentioning

confidence: 99%

Enzymology and Structure of the GH13_31 Glucan 1,6-α-Glucosidase That Confers Isomaltooligosaccharide Utilization in the Probiotic Lactobacillus acidophilus NCFM

et al. 2012

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“…Serum urea nitrogen (SUN) and serum glucose were determined colorimetrically (Vitros 950, Rocheste, NY, USA). Amino acids were determined by HPLC as described by Barkholt and Jensen (1989) with the exception that A-buffer had pH ¼ 3.17 in order to separate citrulline and glutamic acid. Serum insulin and C-peptide was determined with solid phase, two-site fluoroimmunometric assays (AutoDELFIA, PerkinElmer, Turku, Finland).…”

Section: Methodsmentioning

confidence: 99%

High intakes of milk, but not meat, increase s-insulin and insulin resistance in 8-year-old boys

Hoppe¹,

Mølgaard²,

et al. 2004

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Objective: Our objective was to examine if a high animal protein intake from milk or meat increased s-insulin and insulin resistance in healthy, prepubertal children. A high animal protein intake results in higher serum branched chain amino acids (BCAA; leucine, isoleucine and valine) concentrations, which are suggested to stimulate insulin secretion. Furthermore, milk possesses some postprandial insulinotrophic effect that is not related to its carbohydrate content. Design: A total of 24 8-y-old boys were asked to take 53 g protein as milk or meat daily. At baseline and after 7 days, diet was registered, and insulin, glucose, and amino acids were determined. Insulin resistance and beta cell function were calculated with the homeostasis model assessment. Results: Protein intake increased by 61 and 54% in the milk-and meat-group, respectively. In the milk-group, fasting s-insulin concentrations doubled, which caused the insulin resistance to increase similarly. In the meat-group, there was no increase in insulin and insulin resistance. As the BCAAs increased similarly in both groups, stimulation of insulin secretion through BCAAs is not supported. Conclusions: Our results indicate that a short-term high milk, but not meat, intake increased insulin secretion and resistance. The long-term consequences of this are unknown. The effect of high protein intakes from different sources on glucose-insulin metabolism needs further studying.

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“…After hydrolysis for 20 h at 110°C in vacuo in 6 M HC1, 0.1% phenol, 5 ~ thioglycollic acid the amino acid composition was determined by c ttion-exchange chromatography [27] modified as in [28].…”

Section: Amino Acid Composition Analysmentioning

confidence: 99%

Expression and refolding of a high‐affinity receptor binding domain from rat α₁‐macroglobulin

et al. 1995

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A recombinant version of the receptor binding domain of rat at-macroglobulin (RBDv) consisting of residues 1319-1474 has been expressed in E. coil. Competition experiments with USl-labelled methylamine treated human a2-macroglobulin reveal that the th-macroglobulin-RBDv exhibit the same high affinity for the c~2-macroglobulin receptor as the entire 40 kDa light chain from rat a~-macroglobulin. It is therefore concluded, that all determinants for receptor interaction reside in the C-terminal approx. 150 residues of the t~-macroglobulin subunit.

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Amino acid analysis: Determination of cysteine plus half-cystine in proteins after hydrochloric acid hydrolysis with a disulfide compound as additive

Cited by 326 publications

References 14 publications

Enzymology and Structure of the GH13_31 Glucan 1,6-α-Glucosidase That Confers Isomaltooligosaccharide Utilization in the Probiotic Lactobacillus acidophilus NCFM

Enzymology and Structure of the GH13_31 Glucan 1,6-α-Glucosidase That Confers Isomaltooligosaccharide Utilization in the Probiotic Lactobacillus acidophilus NCFM

High intakes of milk, but not meat, increase s-insulin and insulin resistance in 8-year-old boys

Expression and refolding of a high‐affinity receptor binding domain from rat α₁‐macroglobulin

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Amino acid analysis: Determination of cysteine plus half-cystine in proteins after hydrochloric acid hydrolysis with a disulfide compound as additive

Cited by 326 publications

References 14 publications

Enzymology and Structure of the GH13_31 Glucan 1,6-α-Glucosidase That Confers Isomaltooligosaccharide Utilization in the Probiotic Lactobacillus acidophilus NCFM

Enzymology and Structure of the GH13_31 Glucan 1,6-α-Glucosidase That Confers Isomaltooligosaccharide Utilization in the Probiotic Lactobacillus acidophilus NCFM

High intakes of milk, but not meat, increase s-insulin and insulin resistance in 8-year-old boys

Expression and refolding of a high‐affinity receptor binding domain from rat α1‐macroglobulin

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Expression and refolding of a high‐affinity receptor binding domain from rat α₁‐macroglobulin