Combined Chemical-Enzymatic Assembly of Aminoglycoside Derivatives with N-1-AHB Side Chain
Written by Lilach Chen
L. Chen1, I. Nudelman1, N. M. Llewellyn2, E.Sahraoui2,M. Cherniavsky1, J. B. Spencer2, and T. Baasov1
2-Deoxystreptamine (2-DOS) aminoglycosides are highly potent, broad-spectrum antibiotics. The rapid spread of antibiotic resistance towards this family of antibiotics and their relative toxicity to mammals are critical problems that largely limit the intensive clinical use of these drugs.
To overcome these problems, a wide variety of aminoglycoside derivatives along with various modification methods have been documented during the last few decades. Among these methods, a selective N-acylation of the 2-DOS ring (ring II) at the N-1 position with the (S)-4-amino-2-hydroxybutanoyl (AHB) group, turned out to be one of the most successful modifications of natural aminoglycosides, which resulted in derivatives with improved antibacterial activity and reduced acute toxicity relative to that of the parent molecule.
Additionaly, in the last several years, aminoglycosides showed a unique ability to induce mammalian ribosomes to readthrough disease-causing premature nonsense mutations and generate full-length functional proteins. However, the high toxicity of clinical aminoglycosides, along with their reduced readthrough activity at subtoxic doses, prevents their clinical use. Earlier studies demonstrated that the selective addition of AHB moiety at N-1 position leads to the improvement of stop codon readthrough potential along with decrease in toxicity of the tested compounds.
In general, regioselective attachment of the AHB moiety on the aminoglycoside structure frequently requires long protection schemes and the efficiency of a particular strategy is generally dependent on the structure of the parent aminoglycoside. Therefore, we wanted to probe a shorter, enzymatic approach, as an alternative method.
In this study we show a selective acylation at the N-1 position of a series of unprotected pseudo-disaccharides and pseudo trisaccharides of 2-deoxystreptamine-containing aminoglycosides with the AHB pharmacophore by using the recombinant BtrH and BtrG enzymes involved in biosynthesis of butirosin from Ribostamycin, in combination with a synthetic acyl donor. The process was optimized by performing two enzymatic steps in a sequential manner without purification of the intermediate product.