Hydrolyzable amino acids

If specific amino acid-type labeling is required, the labeled amino acid is added to the fermentation of the expression host (topic 1 above, see Sect. 1.2.3). In this case, a thorough isotope analysis of the expressed protein is advisable prior to NMR spectroscopic investigations. This is preferentially achieved by GC-MS analyses of the hydrolyzed amino acids from the protein product. 

Henrichs, S.M., J.W. Farrington, and C. Lee. 1984. Peru upwelling region sediments near 15°S - II. Dissolved free and total hydrolyzable amino acids. Limnology and Oceanography 29 20-34. 

A particularly interesting case is the partial hydrolysis of the racemic N-tri-fluoroacetyl derivative 59 with hog kidney aminoacylase (HKA) (EC 3.5.1.14) to prepare chiral isomers of 2-trifluoromethylalanine 60 [53]. The stereochemical preference of hog kidney aminoacylase is to hydrolyze amino acid amides bearing the larger C-2 substituent in the pro-S position [54]. The enzymatic hydrolysis of compound 60 follows this trend. 

Yield and % ee in parentheses are using the pseudoenantiomeric catalysts 7b or 7a. h%Yield to hydrolyzed amino acid product. cNo Product Obtained. tBuOiCCHjBr. 

The hydrolysis of the acyl-enzyme occurs through a similar process where a proton would be abstracted from the entering nucleophile (H O) by His-57 which is in the out-position (Fig. 2). This allows the formation of a tetrahedral intermediate (26 27). His-57 then moves into the in-position (27 28) and delivers a proton to Ser-195. This permits the breakdown of the intermediate to give the free enzyme 29 and the hydrolyzed amino acid residue 30. 

Note Humic substances (HS) are operationally defined as DOC that is retained on XAD-2 or XAD-8 resins. UDOM is operationally defined as DOC that is retained by a membrane with a 1 nm pore size and 1000 Da molecular weight cutoff. 13C-NMR spectroscopy was used for analysis of carbon functional groups, which are presented as a percentage of the total organic carbon in the sample. THNS, total hydrolyzable neutral sugars THAA, total hydrolyzable amino acids. 

TABLE II The Percentages of Bioavailable DOC (% BDOC) Accounted for as Total Hydrolyzable Neutral Sugars (THNS) and Total Hydrolyzable Amino Acids (THAA) in a Lake, Creek, and Marine Environment... 

Figure 9.34 Total hydrolyzable amino acid (THAA) concentrations (mg L-1) during (a) oxic and (b) anoxic decay of the dinoflagellate P minimum over different time periods. Hydroxylamino acids not shown, and nonprotein amino acids = /3-alanine and y-aminobutync acid. (Modified from Nguyen and Harvey, 1997.)...

The enzyme carboxypeptidase A hydrolyzes amino acids from the C-terminal end of peptides, provided that the C-terminal residue is not proline, lysine, or arginine. Fig. 4-13 shows the sequential release of amino acids from a protein by means of treatment with carboxypeptidase A. Deduce the sequence at the C terminus. 

Figure 15.17 Ratio of basic amino acids (B), (lysine and arginine) to the sum of basic plus acidic amino acids, (B+A), (aspartic and glutamic acids) versus carbon-normalized yields of total hydrolyzable amino acids (THAA). Symbols are as in Fig. 15.13.

Henrichs, S. M., Farrington, J. W., and Lee, C. (1984). Peru upweUing sediments near 15S. Part 2. Dissolved free and total hydrolyzable amino acids. Limnol. Oceanogr. 29, 20—34. 

Several methods for load determination have been described in the literature however, it is important to point out that application of more than one method very often leads to different results.For those resins where incorporation of an amino acid is easily achieved, amino acid analysis of the acid-hydrolyzed amino acid (peptide) resins should be the method of choice because it gives the real amino acid content. The amino acid to be incorporated into the resin should be stable to the acid conditions. Thus, Ser, Thr, Cys, and Tyr should be avoided. After incorporation of the amino acid, washing of the peptide-resin is advisable in order to remove the excess protected amino acid that is not covalently attached to the resin. 

Thurman (1985) provided a comprehensive assessment of the distribution of identifiable organic compounds in natural waters. Total hydrolyzable amino acids accounted for 2-3% of DOC in rivers and 3 -13 % of DOC in eutrophic lakes. Total hydrolyzable sugars accounted for 5-10% of DOC in rivers and 8-12% of DOC in lakes. Thurman (1985) also summarized the... 

The analysis of amino acids can be conducted on unmodified samples to quantify free amino acids, or total hydrolyzable amino acids can be quantified by conducting the analysis on samples that have been hydrolyzed for 24 h in 6 M HCl at 100 °C. In either case, amino acids must be derivatized prior to quantitative analysis. Derivatization is used to render amino acids less... 

Total hydrolyzed amino acids (THAAs) and total free amino acids (TFAAs). 

The major constituents of biomass are present at low levels in DOM. The median freshwater contains 1.3p,molL of total hydrolyzable amino acids, the most abundant of which are glycine (20%), aspartic acid (12%), alanine (10%), glutamic acid (9%), and serine (9%). Collectively, the amino acids account for 1.8% of the organic carbon in DOM and —18% of its nitrogen. Free amino acids also exist in freshwaters, but the median concentration of 0.3 p,mol is a factor of 4-5 lower than the concentration of bound amino acids. Elevated levels of serine and ornithine in the data set for free amino acids strongly suggest that some samples may have been contaminated by human fingerprints. 

Yields (%) are calculated based on input N2 or CO2 (for HCHO) from 100 mm CO2, 100 mm N2 and 100 mL H2O with or without 2 mmoles CaCCb in a 3.1 L flask at 23 C. Also included is an experiment with 10 mm of O2 added to the gas mixture. The results with a reducing atmosphere (% yields based on starting carbon in the form of methane) are given for comparison [taken from (3, 4). Yields of THAA (Total Hydrolyzable Amino Acids labeled in plate 2) are shown after hydrolysis in the absence (-) or presence (+) of ascorbate. The controls are not sparked and represent blanks. ND = not determined. 

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