Deoxypolypeptides bind and cleave RNA

Abstract

We have prepared L-and D-deoxypolypeptides (DOPPs) by selective reduction of appropriately protected polyhistidines with borane, reducing the carbonyl groups to methylenes. The result is a chiral polyamine, not amide, with a mainly protonated backbone and chirally mounted imidazolylmethylene side chains that are mostly unprotonated at neutrality because of the nearby polycationic backbone. We found that, in contrast with the D-octahistidine DOPP, the L-octahistidine DOPP is able to cooperatively bind to a D-polyuridylic acid RNA; this is consistent with results of previous studies showing that, relative to D-histidine, L-histidine is able to more strongly bind to RNA. The L-DOPP was also a better catalyst for cleaving the RNA than the D-DOPP, consistent with evidence that the L-DOPP uses its imidazole groups for catalysis, in addition to the backbone cations, but the D-DOPP does not use the imidazoles. The L-DOPP bifunctional process probably forms a phosphorane intermediate. This is a mechanism we have proposed for models of ribonuclease cleavage and for the ribonuclease A enzyme itself, based on our studies of the cleavage and isomerization of UpU catalyzed by imidazole buffers as well as other relevant studies. This mechanism contrasts with earlier, generally accepted ribonuclease cleavage mechanisms where the proton donor coordinates with the oxygen of the leaving group as the 2-hydroxyl of ribose attacks the unprotonated phosphate. T he enzyme ribonuclease A (RNase A) hydrolyzes RNA with bifunctional catalysis, using His-12 as a base and protonated His-119 as an acid to form the first products, a 2,3-cyclic phosphate of one ribose and the free next ribose with a new terminal 5′-hydroxyl group (1, 2). Lys-41 is also involved in stabilizing intermediate anions (3). In a second step the cyclic phosphate is hydrolyzed using the now-protonated His-12 and now-deprotonated His-119 in essentially a reverse of the cyclization step, with water replacing the second ribose unit (4). In the classical mechanism of this process it had been generally accepted, including in textbooks, that the protonated His-119 donates its proton to the leaving group whereas the 2′-hydroxyl group attacks the phosphate (1, 2) [isotope studies show that two protons are "in flight" during the hydrolysis step, essentially the reverse of the cyclization, both moving at the same time (5)]. We also showed the two-proton-in-flight process in a model system (6).However, our extensive studies on the cleavage of RNA with imidazole-imidazolium catalysts and on catalysis by RNase A itself indicated that a different mechanism is used ( Fig. 1) (7, 8). The proton donor BH + hydrogen binds to a phosphate oxyanion and donates the proton to the phosphate as the ribose 2′-hydroxyl adds while losing a proton to base :B. This process results in an intermediate phosphorane species. This then expels the 5′-hydroxyl group of the second nucleotide to cleave the P-O bond in a second step. We proposed this two-step phosphorane process to be the preferred path for th...

Chiral Discrimination of Diamines by a Binaphthalene-Bridged Porphyrin Dimer

“…Repeated chromatography followed by recrystallization from chloroform and methanol gave pure (R)-H1 as a purple powder (12 mg, 20%). 1 4.2. Synthesis of (S)-H1.…”

“…Chiral discrimination originates from the different physical and chemical properties of diastereomeric supramolecular systems, which is generally involved in the supramolecular interactions of chiral species. The related effects hold very important functions in biological systems , and have many practical applications in absolute configuration (AC) assignment, enantiomer resolution, enantiomeric excess (ee) analysis, molecular recognition, , asymmetric catalysis, and materials science. , Consequently, accurate and efficient chiral discrimination systems are highly desirable.…”

Magnesium pyrophosphates in enzyme mimics of nucleotide synthases and kinases and in their prebiotic chemistry

“…A good example from our own work includes the mechanisms used when thiamine pyrophosphate is the coenzyme; thiamine itself can perform the essential catalysis, but more slowly and less selectively than with the enzyme (1, 2). As another example, our mimics of ribonuclease use the imidazole groups that are biochemically part of the enzyme itself to catalyze cleavage of RNA (3,4). We here describe mimics of the enzymes that attach base groups to a ribose unit.…”