Lysine, labeled

Figure 4. HSQC spectra of specifically N-lysine labeled apoLp-III in the presence Oight...

Lysine-labelled tryptophanase was prepared by Incubating the enzyme with 100-fold molar excess of 2,2,5,5-tetramethyl-3-pyrrolin-1-oxyl-3-carboxylic acid N-hydroxysuccinimide ester (Eastman Kodak Company) in 10 mM bis-tris-propane (Sigma), pH 8 (20% v/v... 

The effect of wQ on the enzyme was examined by comparing the EPR spectra of lysine-labeled tryptophanase. Although the spin label is nonspecific, it can still provide molecular-level information about the enzyme under different conditions. EPR spectra of the spin-labeled enzyme in bulk water and in reverse micelles are shown in Figure 6. Much broader spectra were obtained in reverse micelles, and the calculated rotational correlation time of the attached label (101 increased with w0. Thus, the enzyme-bound spin label became more constrained as the water content of the reverse micelle decreased. Since the rotational correlation time of the entire protein in bulk water calculated from the Stokes-Einstein equation is about 200 nsec, the motion of the spin label was still rapid relative to the tumbling rate of the enzyme. Therefore, broadening of the spectrum was apparently caused by a change in the local dynamics of tryptophanase rather than by a decrease in the enzyme s overall rotation rate. The tumbling rate of the enzyme could have decreased as well, however. 

EPR sprectra of lysine-labeled tryptophanase in (A) bulk water and (B) reverse micelles. 

An ingenious model has been developed which accounts for the biosynthesis of all piperidine alkaloids (cf. Vol. 10, p.9). The key idea is that those alkaloids which are formed with symmetriz-ation of a lysine label are biosynthesized by way of free cadaverine (17) those which are formed without symmetrization of label are biosynthesized by way of cadaverine,which remains unsymraetrical by being (co)enzyme-bound [as (15)] until oxidation occurs to give (16) (Scheme 2). 

Slaframine.—Further details are now available on the biosynthesis of slaframine (39), a toxin produced by the mould Rhizoctonia leguminicola. Both dl-[1- C]-and DL-[6- " C]-lysine afforded labelled slaframine, indicating intact incorporation of all the carbons of the amino-acid. Addition of inactive pipecolic acid (34) to the cultures diluted the lysine label in the derived slaframine, and pipecolic acid was labelled by radioactive lysine. Further carboxyl- and ring-labelled pipecolic acids [as (34)] were both well incorporated into slaframine. A clear indication is thus obtained of the biosynthetic sequence lysine —> pipecolic acid (34) — slaframine (39) 2-hydroxymethylpiperidine is not a precursor. Attention is drawn to the discovery of a similar pathway to a metabolite of similar structure also produced by R. leguminicola. ... 

Proton relaxation enhancement by means of serum albumin and poly-lysine labeled with DTPA-Gd relaxivities as a function of molecular weight and conjugation efficiency. Magn. Reson. Imaging 10(6) 913-917... 

LaBadie, J., W.A. Dunn, and N.N. Jr. Aronson. 1976. Hepatic synthesis of carnitine from protein-bound trimethyl-lysine. Lysosomal digestion of methyl-lysine-labelled asialo-... 

Treatment of the heptapeptide with 2,4-dinitrofluorobenzene followed by incomplete hydrolysis gave, among other products valine labeled at the a-amino group, lysine labeled at the s-amino group, and a dipeptide, DNP—VL (DNP = 2,4-dinitrophenyl-). 

The above example is very simple where the method of conventional organic synthesis did not need to be modified much. There are. however, other labeled s)mtheses where the organic synthesis method has to be modified profoimdly. One such example is the mthesis of DL-lysine labeled in the e-carbon atom. One interesting problem arises here. Since this synthesis is a multistep process, the isotope can be introduced either at an early step in the experiment, or in a later step. The latter option seems ro firom the point of view of conserving the radioactive isotope. However, it is experimentally found that introduction of isotope at a later step in the experiment does not yield the desired product. Therefore, even though the yield would be less, it is necessary that liie isotope be introduced at an early step in the mthesis. The reaction steps are givert low. 

Two bromoacetyl derivatives of DNP ligands of different length (BADE and BADL) were shown to specifically label either light or heavy chains. BADE (o-A-bromoacetyl-y-A-DNP-ay-diamino-L-buty-ric acid) labels Tyr-34 on the light chain of protein 315, whereas BADL (a-A-bromoacetyl- -iV-DNP-L-lysine) labels Lys-54 on the heavy chain of protein 315. ... 

The results of extensive experiments with variously labelled samples of lysine closely define the way in which this amino acid is assimilated into the alkaloids. Thus C-2 and C-6 of the precursor become C-2 and C-6, respectively, in 6.19) [9], 6.21) [10, 11], and 6.20) [12]. Although cadaverine 6.26) is also an alkaloid precursor it cannot be an intermediate following lysine because any label at C-2 or C-6 of the amino acid would become spread over both C-1 and C-5 of the symmetrical diamine 6.26) a single lysine label would thus be spread over both C-2 and C-6 of the alkaloids. 

Scheme 2 Preparation of lysine-labeling probes. DOTA-OSu, EtsN, DMF, rt, 58% coumarin-OSn, CH2CI2, rt, 68% TAMRA-OSu, DMF/CH2CI2 (1 1), rt, 74% Cy5-0Su, DMF/CH2CI2 (1 1), rt, 60% Dess-Martin periodinane, DMF/CH2CI2 (3 10), rt or polystyrene-supported IBX, DMF/CH2CI2 (1 1), It...

Lysine labeling engineering kit STELLA from Kishida Chemical Co., Ltd., http //www. kishida.co.jp/... 

Taking into account the fundamental clinical problem of our work, we have chosen the amino acid lysine, labelled with (Fig 1)... 

This experiment shows that valine is the N-terminal amino acid and that valine is attached to leucine. (Lysine labeled at the e-amino gronp is to be expected if lysine is not the N-terminal amino acid and if it is linked in the polypeptide through its a-amino group.)... 

LaBadie, J. Dunn, W.A. Aronson, N.N. Jr. Hepatic Synthesis of Carnitine from Protein-Bound Trimethyl-Lysine. Lysosomal Digestion of Methyl-Lysine-Labelled Asialo-Fetuin. Bio-chem. J. 1976, 160, 85-95. 

Dunn, W.A. Englard, S. Carnitine Biosynthesis by the Perfused Rat Liver from Exogenous Protein-Bound Trimethyllysine. Metabolism of Methylated Lysine Derivatives Arising from the Degradation of 6-N-[Methyl- H] Lysine-Labeled Glycoproteins. J. Biol Chem. 1981, 256, 12437-12444. 

In addition to nonspecific lysine labeling. Lien et al. reported site-specific incorporation of a fluorescent tag into nascent proteins using a Cys-tRNA . After aminoacylation of E. coli Cys-tRNA , it was modified with a SH-reactive fluorophore (BODIPY-FL). This method may prove valuable when used in conjunction with site-directed mutagenesis to create protein containing single-cysteine residues by which one can prepare the nascent protein containing single fluorophore at a defined position. 

These results are consistent with the formation of ring B from lysine (labelled as indicated on the next page from [1,2- C2]-acetate). The lysine then affords plpecollc acid (1). Chain extension then takes place with an acetate unit and a three-carbon unit (possibly from the glycolytic pathway). )... 

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