Acceptor Specificity

Interestingly, the E. coli enzyme s relaxed acceptor specificity allows for substitution of both cosubstrates, albeit at strongly reduced (<1% of v, catalytic rates. Propanal, acetone, or fluoroacetone can replace ethanal as the donor in the synthesis of variously substituted 3-hydroxyketones such as (112) or (113) (Figure 10.41)... 

In a very recent approach Seibel et al. used a microarray approach on microtiter plates to identify new acceptor specificities of the non-Leloir GTFR from S. oralis,... 

Irradiation at >320 nm releases from the polymer, whether insoluble or water-soluble, free oligosaccharides in very high yields. A simple illustration of such a sequence carried out with either insoluble 2-aminoethyl-substituted poly(acrylamide) beads or with water-soluble, substituted poly(vinyl alcohol) is presented in Scheme 9 the isolated overall yield of lactose was 29.9% (soluble-polymer approach). The synthesis on light-sensitive polymers facilitates the isolation of products, which is important from the preparative point of view and as a tool for the study of enzymes, permitting efficient comparison of acceptor specificity and being capable of demonstrating de novo synthesis. 

Different GTases have different acceptor specificities ( ). 

Table I. Acceptor Specificity of Several Partially Purified Glucoayl-...

Substrate-specificity studies on microsomal, frog-liver sialyltrans-ferase revealed the presence of (2—>3) and (2—>6) activities.277 This enzyme system readily sialylates oligosaccharides, but is almost inactive with asialofetuin, which is in contrast to the sialylation of oligosaccharides, as well as asialofetuin, by rat-liver sialyltransferase.278 The conclusion from this observation is that acceptor specificity of sialyl-transferases isolated from liver of evolutionary distant animals is similar for substrates of low molecular weight, but differs for compounds of high molecular weight.279... 

G. F. Herrmann, P. Wang, G.-J. Shen, E. Garcia-Juneeda, S. H. Khan, K. L. Malta, and C.-H. Wong, Large-scale production of recombinant a-1,2-mannosyltranferase from E. coli for the study of acceptor specificity and use of the recombinant whole cells in synthesis, J. Org. Chem. 59 6356 (1994). 

Discrimination between some pairs of tRNAs depends entirely on the anticodon sequence. For example, tRNAMet contains the anticodon CAU. That for a minor tRNAIle is the same except that the cytosine has been posttranscriptionally modified by covalent linkage of a molecule of lysine via its e-amino group to C2 of the cytosine. The latter base (Iysidine) is correctly recognized by E. coli isoleucyl-tRNA synthetase but, if the cytosine is unmodified, it is aminoacylated by methionyl-tRNA synthetase.192 In most instances the acceptor specificity, or tRNA identity, is not determined solely by the anticodon sequence. Thus, when a methionine initiator tRNA was modified to contain a tryptophan anticodon, it was only partially charged with tryptophan in vivo. However, when A73 of the methionine tRNA was also converted to G73, only tryptophan was inserted.193 Nucleotide 73 (Fig. 

Since the enzyme must also catalyze the reverse reaction the acceptor specificity should extend to all compounds of the type R—XH. 

Functionally and mechanistically reminiscent of the pyruvate lyases, the 2-deoxy-D-ribose 5-phosphate (121) aldolase (RibA EC 4.1.2.4) [363] is involved in the deoxynucleotide metabolism where it catalyzes the addition of acetaldehyde (122) to D-glyceraldehyde 3-phosphate (12) via the transient formation of a lysine Schiff base intermediate (class I). Hence, it is a unique aldolase in that it uses two aldehydic substrates both as the aldol donor and acceptor components. RibA enzymes from several microbial and animal sources have been purified [363-365], and those from Lactobacillus plantarum and E. coli could be induced to crystallization [365-367]. In addition, the E. coli RibA has been cloned [368] and overexpressed. It has a usefully high specific activity [369] of 58 Umg-1 and high affinity for acetaldehyde as the natural aldol donor component (Km = 1.7 mM) [370]. The equilibrium constant for the formation of 121 of 2 x 10M does not strongly favor synthesis. Interestingly, the enzyme s relaxed acceptor specificity allows for substitution of both cosubstrates propional-dehyde 111, acetone 123, or fluoroacetone 124 can replace 122 as the donor [370,371], and a number of aldehydes up to a chain length of 4 non-hydrogen atoms are tolerated as the acceptor moiety (Table 6). 

Harper, M., Boyce, J.D., Cox, A.D., St. Michael, F., Wilkie, I.W., Blackall, P.J., Adler, B., Pasteurella multocida espresses two lipopolysaccharide glycoforms simultaneously, but only a single form is required for virulence identification of two acceptor-specific heptosyl I transferases. Infect Immun 75 (2007) 3885-3893. 

Kaniuk, N.A., Vinogradov, E., Whitfield, C. Investigation of the structural requirements in the lipopolysaccharide core acceptor for ligation of O antigens in the genus Salmonella WaaL ligase is not the sole determinant of acceptor specificity. J Biol Chem 279 (2004) 36470-36480. 

Seibel J, Hellmuth H, Hofer B, Kicinska AM, Schmalbruch B (2006) Identification of new acceptor specificities of glycosyltransferase R with the aid of substrate microarrays. Chem-biochem 7 310-320... 

Donor Acceptor Specific activity Inhibition by Amytal, rotenone... 

Cytochrome 62 is stereospecific for l(- -)-lactate. It also oxidizes other a-hydroxymonocarboxylic acids at slow rates 80, 96). As electron acceptors ferricyanide, methylene blue, 2,6-dichloroindophenol, 1,2-naphthoquinone 4-sulfonate, and cytochrome c have been used. This wide acceptor specificity is characteristic of a number of flavoproteins, which are generally capable of reducing quinoid structures and ferric compounds 97). However, as will be seen below, cytochrome c is considered to be the physiological electron acceptor for the yeast L-lactate dehydrogenase. 

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