Threonine synthetase mechanism

In a study of the mechanism of the synthetase reaction Flavin and co-workers determined the incorporation of 0 (108) and of deuterium from the water (109) of the incubation medium into the products of the reaction. It was found that one atom of solvent oxygen was incorporated into threonine, and none into phosphate. This shows that the latter is removed by nonhydrolytic elimination, rather than hydrolysis. Incubation of 0-phos-phohomoserine with purified threonine synthetase in 100% DjO resulted in the incorporation of two atoms of D in the newly formed threonine, one in the a-position, the second in the -y-position. 

Fio. 4, Probable mechanism of threonine synthetase in formation of threonine. Modified from Flavin and Slaughter (f09). Experiments with HiO and DjO were performed independently. Pyridine substituents have been omitted from intermediates for convenience. 

The zinc site is less well suited to discrimination against serine because this amino acid does have a hydroxyl group that can bind to the zinc. Indeed, with only this mechanism available, threonyl-tRNA synthetase does mistakenly couple serine to threonyl-tRNA at a rate 10 2 to 10 3 times that for threonine. As noted in Section 29. LL this error rate is likely to lead to many translation errors. How is a higher level of specificity achieved ... 

Tiessel, T, Thompson, R., Zieske, L. R., Menendez, M., and Davis, L. (1986). Interaction between L-threonine dehydrogenase and aminoacetone synthetase and mechanisms of artiinuacetone production. /. Bjuf. Citon. 261,16428-16437. 

Coordination of the threonine hydroxyl by an active site Zn in the threonyl-tRNA synthetase allows discrimination between threonine and the isosteric valine (Sankaranarayanan et al., Nat. Struct. Biol. 7[2000] 461-465). Given the similarity of serine and threonine (Ser lacks only the methyl group of Thr), if this is the only mechanism for amino acid discrimination available, threonyl-tRNA synthetase mistakenly couples Ser to threonyl-tRNA at a rate several-fold higher than it does threonine. Since this would lead to unacceptably high error rates in translation, how it is it avoided ... 

In other words, isoleucine and valine stabilize the enzyme in the immature form. Their further observation that the immature form of threonine deaminase has an affinity for leucyl tRNA but not for valyl or isoleucyl tRNA is certainly of interest with respect to the obvious correlation with multivalent repression that is possible [16]. Hatfield and Burns propose that the leucyl tRNA-immature tetramer complex might be the holorepressor of the ilv genes or perhaps at least of the ilv ADE operon. If either valine or isoleucine were limiting, maturation of the enzyme would occur without interference by either leucine or leucyl tRNA, thus leading to derepression. Derepression would also occur when isoleucine and valine were both in sufficient excess to block maturation provided leucine were limiting so that the leucyl tRNA-immature tetramer complex could not be formed. The significance of this model has yet to be assessed, but it is difficult at present to account for the supposed involvement of the isoleucyl and valyl tRNA synthetases in multivalent repression. Nevertheless, this interesting observation should be kept in mind as a possible mechanism that could account for part of the phenomenon of multivalent repression when additional information is acquired. 

---