β-Hydroxylation of α-amino-β-hydroxylbutanoyl-glycyluridine catalyzed by a nonheme hydroxylase ensures the maturation of caprazamycin

Abstract

Caprazamycin is a nucleoside antibiotic that inhibits phospho-N-acetylmuramyl-pentapeptide translocase (MraY). The biosynthesis of nucleoside antibiotics has been studied but is still far from completion. The present study characterized enzymes Cpz10, Cpz15, Cpz27, Mur17, Mur23 out of caprazamycin/muraymycin biosynthetic gene cluster, particularly the nonheme αKG-dependent enzyme Cpz10. Cpz15 is a β-hydroxylase converting uridine mono-phosphate to uridine 5′ aldehyde, then incorporating with threonine by Mur17 (Cpz14) to form 5′-C-glycyluridine. Cpz10 hydroxylates synthetic 11 to 12 in vitro. Major product 13 derived from mutant Δcpz10 is phosphorylated by Cpz27. β-Hydroxylation of 11 by Cpz10 permits the maturation of caprazamycin, but decarboxylation of 11 by Mur23 oriented to muraymycin formation. Cpz10 recruits two iron atoms to activate dioxygen with regio-/stereo-specificity and commit electron/charge transfer, respectively. The chemo-physical interrogations should greatly advance our understanding of caprazamycin biosynthesis, which is conducive to pathway/protein engineering for developing more effective nucleoside antibiotics.

Naa10 regulates hippocampal neurite outgrowthviaBtbd3 N-α-acetylation-mediated actin dynamics

“…Briefly, the gradient employed was 2% buffer B at 2 min to 40% buffer B at 40 min. Under full-scan MS condition, the mass range was m/z 375-1800 (AGC target 5E5) with a lock mass, resolution of 60,000 at m/z 200, and a maximum injection time of 50 ms. 54 Target m/z values were isolated for CID with NCE of 35 and a maximum injection time of 100 ms. The electrospray voltage was maintained at 1.8 kV, and the capillary temperature was set at 275 °C.…”

Decarboxylation in Natural Products Biosynthesis

“…It is worth noting that LolD is not the only type III PLP-dependent decarboxylase that can process substrates other than common amino acids. For example, the enzyme Mur23, found in the muraymycin biosynthetic pathway, can decarboxylate nucleoside-containing substrates (Figure B). , Additionally, type III PLP-dependent decarboxylases can also process larger substrates such as peptides and amino acids tethered to carrier proteins in RiPP and NRP biosynthesis, respectively. These findings highlight the versatility and potential of type III PLP-dependent decarboxylases as promising biocatalysts in various biosynthetic pathways and suggests that much remains to be discovered in the substrate range of this enzyme class.…”

Intermolecular Interactions of Nucleoside Antibiotic Tunicamycin with On-Target MraYCB-TUN and Off-Target DPAGT1-TUN in the Active Sites Delineated by Quantum Mechanics/Molecular Mechanics Calculations

“… 30 − 32 Bader’s quantum theory of atoms in molecules (QTAIM) 33 − 35 and natural bond orbital (NBO) analyses 36 , 37 are two reliable theoretical approaches to gain further insights into the nature and strength of the intermolecular interactions, especially the H-bonding interactions, in the protein structures. 24 , 25 , 38 − 40 …”

“…From a theoretical standpoint, comprehensive description of these interactions requires the use of applicable computational methods. Nowadays, parallel progress in computational softwares and development in computer architectures, the combined quantum mechanics/molecular mechanics calculations have been deemed as a powerful computational chemistry method to study the protein–ligand interactions. − On the other hand, the M06 family of density functional theory (DFT) comprises suitable functionals to compute noncovalent intermolecular interactions, such as hybrid meta-GGA density functional (M06-2X). − Bader’s quantum theory of atoms in molecules (QTAIM) − and natural bond orbital (NBO) analyses , are two reliable theoretical approaches to gain further insights into the nature and strength of the intermolecular interactions, especially the H-bonding interactions, in the protein structures. ,,− …”