Threonine isolation

C-2 and C-3 are each bonded to four different substituents. Therefore, threonine has two chiral centers. Because the chiral centers are nonequivalent, four diastereomers are possible. The Fischer projections are written by placing the carboxyl group at the top of the vertical chain. The amino and hydroxyl groups can be on the right or left sides of the projection formula. The structure of threonine isolated from proteins is given by the Fischer projection at the right. Its configuration is 2S,3R. 

Amino acids are the main components of proteins. Approximately twenty amino acids are common constituents of proteins (1) and are called protein amino acids, or primary protein amino acids because they are found in proteins as they emerge from the ribosome in the translation process of protein synthesis (2), or natural amino acids. In 1820 the simplest amino acid, glycine, was isolated from gelatin (3) the most recendy isolated, of nutritional importance, is L-threonine which was found (4) in 1935 to be a growth factor of rats. The history of the discoveries of the amino acids has been reviewed... 

Sirolimus (SRL), also termed rapamycin is a macrolide lactone isolated from the ascomycete species Stre-ptomyces hygroscopicus. After binding to its cytosolic receptor FKBP-12 the resulting complex inhibits the multifunctional serine-threonine kinase mTOR (mammalian target of rapamycin). Inhibition of mTOR prevents activation of the p70S6 kinase and successive... 

Removal of the oligosaccharide chain from the glycoprotein usually involves the use of one of three main methods treatment with NaOH-NaBH4, hydrazinolysis, or proteolysis.34-36 The NaOH-NaBH4 treatment is used to release, somewhat specifically, oligosaccharides O-linked to serine and threonine. Hydrazinolysis is used to break N-linkages, and proteolysis, to isolate glycopeptides. Each method apparently still has some drawbacks. 

A murine male germ cell-assocated kinase (mak) was isolated by virtue of its weak homology to the v-ros tyrosine kinase (Matsushime et al., 1990). Although it was isolated because of its homology to a tyrosine kinase, sequence analysis revealed that it belongs to the serine/threonine... 

The second method also relies on site-specific chemical modification ofphosphoproteins (Oda et al., 2001). It involves the chemical replacement of phosphates on serine and threonine residues with a biotin affinity tag (Fig. 2.7B). The replacement reaction takes advantage of the fact that the phosphate moiety on phosphoserine and phosphothreonine undergoes -elimination under alkaline conditions to form a group that reacts with nucleophiles such as ethanedithiol. The resulting free sulfydryls can then be coupled to biotin to create the affinity tag (Oda et al., 2001). The biotin tag is used to purify the proteins subsequent to proteolytic digestion. The biotinylated peptides are isolated by an additional affinity purification step and are then analyzed by mass spectrometry (Oda et al., 2001). This method was also tested with phosphorylated (Teasein and shown to efficiently enrich phosphopeptides. In addition, the method was used on a crude protein lysate from yeast and phosphorylated ovalbumin was detected. Thus, as with the method of Zhou et al. (2001), additional fractionation steps will be required to detect low abundance phosphoproteins. 

Serine/threonine kinase activity has been reported in ESP of pepsin-HCl isolated muscle larvae (Arden el al., 1997) and kinase activity was associated with proteins of 70 and 135 kDa. Phosphorylation status is functionally significant for multiple regulatory factors, including those involved in muscle differentiation (Li et al., 1992). Therefore, kinase activity in parasite secretions may be significant in either the muscle or intestinal phases of infection. 

Fig. 1. Modification of plant metabolic pathways for the synthesis of poly(3HB) and poly(3HB-co-3HV). The pathways created or enhanced by the expression of transgenes are highlighted in bold, while endogenous plant pathways are in plain letters. The various transgenes expressed in plants are indicated in italics. The ilvA gene encodes a threonine deaminase from E. coli. The phaARe, phaBRe, and phaCRe genes encode a 3-ketothiolase, an aceto-acetyl-CoA reductase, and a PHA synthase from R. eutropha, respectively. The btkBRe gene encodes a second 3-ketothiolase isolated from R. eutropha which shows high affinity for both propionyl-CoA and acetyl-CoA [40]. PDC refers to the endogenous plant pyruvate dehydrogenase complex...

By means of a procedure described above, Hanson and Fittkau (HI) isolated seventeen different peptides from normal urine. One of them, not belonging to the main peptide fraction, consisted of glutamic acid, and phenylalanine with alanine as the third not definitely established component. The remaining peptides contained five to ten different amino acid residues and some unidentified ninhydrin-positive constituents. Four amino acids, i.e., glutamic acid, aspartic acid, glycine, and alanine, were found in the majority of the peptides analyzed. Twelve peptides contained lysine and eight valine. Less frequently encountered were serine, threonine, tyrosine, leucine, phenylalanine, proline, hydroxyproline, and a-aminobutyric acid (found only in two cases). The amino acid composi-... 

Characteristically, legume seeds are rich in protein and contain intermediate to high levels of lysine and threonine which are important in balancing the deficiencies of these essential amino acids in cereal diets. Certain legume proteins, such as soybean, also exhibit strong functional properties, especially water solubility, water and fat binding and emulsification. Thus soybean flours, protein concentrates and isolates have been used widely as nutritional supplements and functional ingredients in foods. 

Phosphatidyl serines (or threonine) nh3 —ch2ch.cooh Widely distributed but in small amounts. Is a major component of brain and red cell lipids Usually isolated as salts with K+, Ca2+, Na+ or Mg2+... 

A variety of domain or motif families occur only as extensions to other domains. The Bruton s tyrosine kinase motif (BTK), for example, is found only at the C terminus of PH domains. Similarly, a C-terminal extension (the S TK X domain) to some subfamilies of serine/threonine kinases (S TK) is not found in isolation. Cases where only the extension, and not the preceding domain, is found are strong evidence that the proteins are wrongly assembled from genomic sequence or else represent partial cDNA sequences (Fig. 9, see Color insert). Indeed, all five proteins annotated in SMART as containing a S TK X domain with no catalytic domain are noted to be fragments in their corresponding sequence database entries. 

The complete identification of the amino acids which are essential in the diet is due to W.C. Rose (1938). His first attempts to replace casein with its constituents were unsuccessful because an essential amino acid component in the protein hydrolysate had been missed. After threonine had been isolated by him from casein and fibrin, and shown to be essential, Rose identified val, met, his, lys, phe, leu, ile, thr, and arg as... 

Isolation of individual amino acids started about 1820 by 1904 all of the naturally occurring amino acids in proteins had been isolated except methionine (Mueller, 1922) and threonine (Rose, 1937). One of the earliest methods for the separation of amino acids was through the differential volatility of their methyl or ethyl esters (Emil Fischer, 1901). This approach led to the discovery of valine, proline, and hydroxyproline. [In the 1970s Fischer s method was modified for microanalysis of proteins, separating the amino acid esters by gas phase chromatography. Separation is now usually performed by hplc (high pressure liquid chromatography).]... 

Differently from serine, ESI-MS analysis of homoserine (HSer) solutions reveals an unusually abundant diprotonated homoserine octamers [(HSer)g-2H], but not the expected monoprotonated [(HSer)g H]" one." A 3/1 mixture of L-serine and L-homoserine yields abundant mixed serine octamers with the incorporation of one or two homoserine molecules into the cluster. CID of the isolated [(Ser)6(HSer)2-H] cluster leads to the preferentially loss of two neutral serine molecules. Homoserine is always retained. The ESl-MS spectral patterns of threonine and allothreonine solutions is similar to that of homoserine. A 1/1 mixture of D-serine and D-threonine yields abundant mixed singly- and doubly-charged octamers incorporating from 2 to 6 threonine molecules. Their relative abundance indicates that threonine may incorporate freely into serine clusters because the additional methyl group does not interfere with the bonding of the cluster. 

A number of nonprotein amino acids with unsaturated side chains have been isolated. Many of these contain alkene side chains, but some alkyne side chains containing amino acids have also been identified. Nonprotein dehydroamino acids do not have an a-stereocenter these amino acids are still classified under this category. Dehydroamino acids are generally biosynthesized by the enzymatic elimination of a leaving group at the /3-carbon. For example, serine and threonine are enzymatically dehydrated to give dehydroalanine and dehydrobutyrine, respectively. A similar biosynthetic pathway is hypothesized for dehydroamino acids found in nonribosomal peptides, such as nodularins and microcystins. ... 

Condensation products of DHB (which usually is found also in the fermentation broth) with amino acids were reported, viz. with glycine ixom Bacillus subtilis (164) named subsequently itoic acid (282) with serine from Escherichia coli (261) and Klebsiella oxytoca (196) with threonine from Klebsiella oxytoca (196) and Rhizobium spp. (275, 327) with arginine from Pseudomonas stutzeri (62) with glycine and threonine from Rhizobium sp. (240) with threonine and lysine as well as with leucine and lysine from Azospirillum lipoferum (312, 320). In most cases the isolate (sometimes designated as being a siderophore) was hydrolyzed and the constituents were determined by paper chromatography. The relative amounts of the constituents, the chiralities of the amino acids and the molecular mass of the isolate have not been determined. Hence it is not known whether condensation products of the enterobactin type exist. 

Methanoserine or cleonine 106 has been isolated from cleomycin 107 a, an antibiotic from the bleomycin-phleomycin group, which is different from bleomycin 107b only in its threonine moiety [38]. This amino-(1-hydroxycyclo-propyl)acetic acid is located in the place of the threonine, Eq. (42). 

These two polypeptides have been shown to have identical amino acid composition (Table 9) but to differ from each other in sialic acid content. Da and D4 have 1 and 2 moles of sialic acid per mole of protein, respectively (A5, A6, B9, BIO, E5). A third form without sialic acid has been isolated by preparative isoelectric focusing (A5). Both D3 and D4 have the same NHa-terminal (serine) and COOH-terminal (alanine) amino acid residue and a molecular weight of about 10,000. The complete amino acid sequence has recently been announced (B6) and is reported in Fig. 7. These studies show that the polysaccharide having sialic acid as its terminal sugar, is linked to threonine 74 of the polypeptide chain. 

Finally, recent developments on research into the first C-F bond forming enzyme are summarized. The fluorinase enzyme isolated from Streptomyces cat-tleya catalyzes the formation of 5 -fluoro-5 -deoxyadenosine from S-adenosyl-L-methionine and fluoride. The substrate specificity and subsequent transformation of the fluorinated nucleoside to fluoroacetic acid and to fluoro threonine are discussed. 

Cantharidin is a toxin isolated from fish. Among its numerous biological activities, cantharidin exhibits insecticide and herbicide properties. Cantharidin and norcanthar-idin are inhibitors of serine and threonine phosphatases, which are important enzymes for the regulation of cell processes and cell proliferation. Mono- and difluoronor-... 

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