Mechanism of urea participation in phosphonate ester hydrolysis. Mechanistic and stereochemical criteria for enzymic formation and reaction of phosphorylated biotin

Abstract

The mechanism of participation of a urea group in hydrolysis of a phosphonate ester in acidic solution was investigated. Participation of the oxygen center of the ureido group of 1 and 3 at the phosphonate center is consistent with (1) the high degree of facilitation of ester hydrolysis compared to molecules lacking the adjacent functional group, (2) the observation that species resulting from attack of proximal nitrogen atoms of 1 and 3 are more stable than their precursors, and (3) the observation that acidic, anhydrous media containing alcohols cause rapid transesterification. A detailed mechanistic examination suggests that the participation reaction involves initial formation of a pentacovalent phosphorus intermediate with the ratedetermining step overall being breakdown of the intermediate to expel the leaving alcohol. The potential involvement of Ophosphobiotin in adenosine 5'-triphosphate (ATP) dependent carboxylations in enzymic systems is considered with regard to these results, enzymic results from other laboratories, and stereochemical restrictions. It is concluded that all available experimental evidence is consistent with, but does not require, intermediate formation of O-phosphobiotin. Enzymic carboxylation of O-phosphobiotin, leading to formation of iV-carboxybiotin and phosphate, is considered in terms of the stereochemistry of displacement at the phosphorus center of phosphobiotin. An "adjacent" mechanism, not involving free carboxyphosphate, and an "in-line" mechanism, involving free carboxyphosphate are possible. Reactions not involving O-phosphobiotin are likely to involve "in-line" displacement by bicarbonate at the terminal phosphate residue of ATP, requiring inversion of configuration at phosphorus. Since formation of O-phosphobiotin should occur with inversion at phosphorus, and transfer to bicarbonate with inversion or retention, the route via O-phosphobiotin can account for net inversion or net retention at phosphorus.

Synthesis, structure, and hydrolysis of esters of strained and unstrained N-phosphonylureas

“…Addition of anhydrous ether caused the product to precipitate. 1,4-Dimethyl-2-methoxy-1,4,2-diazaphospholidine-2,5-dione (2) This was prepared by the reaction of N,N1-dimethyl-N-methoxymethylurea and trimethyl phosphite using the method reported by Kluger et al (4).…”

“…This was prepared by the reaction of N,Nf-dimethyl-N-methoxymethylurea and triethyl phosphite (4). Sodium 1,4-dimethyl-2-o.ry-1,4,2-diazaphospholidine-2,5-dione (6) was prepared by reaction of sodium iodide (0.47 g) in 25 mL dry butanone with 2 with stirring and refluxing for 3 h (5).…”

“…The kinetic effects of competition between these and other factors in structurally defined systems provide a quantitative basis for evaluation of the factors controlling reactivity. In this paper we report the rate of hydrolytic reactions of esters of cyclic phosphonylureas (1-3) in alkaline aqueous acetonitrile and the reactions of analogues in which the phosphorus atom is not contained in a ring (4,5). An X-ray crystal study of the cyclic phosphonylurea ester (I) provides a basis for the necessary structural analysis.…”

Carbon‐13 labelled biotin ‐ a new probe for the study of enzyme catalyzed carboxylation and decarboxylation reactions

“…Despite this fact very little is known about the mechanism of carboxylation and COz-transfer within the active sites of these enzymes. 0-Phosphorylated forms of biotin [3,4] as intermediates or attack of a biotin enol at carboxyphosphate have been discussed [5]. A chemically induced carboxylation yielding carboxybiotin in vitro has not been reported.…”

Structure et conformation du thioxo-2 1,3-benzoxazolyl-3 phosphonothioate de diphényle,analogue d'une forme 'N-phosphobiotine'

“…Parmi les hypotheses m~canistiques qui concernent le fonctionnement de la biotine (Lynen, 1967), coenzyme des carboxylases, l'une d'entre elles (Blonski, Belghith, Kla~b~ & P~ri~, 1987) fait intervenir une N-phosphobiotine activ~e par r~action avec l'ad~nosine triphosphate suivant le schema 1. A cette &ape de la r~action deux possibilit~s r&actionnelles sont ~ consid~rer: la premiere implique que la N-phosphobiotine se r6arrange en O-phosphobiotine (phosphate d'&nol) entit6 suffisement r~ac-tive pour jouer le r61e d'activateur de carbonate dans le site enzymatique (Kluger & Adawadkar, 1976;Kluger, Davis & Adawadkar, 1979) …”