Dataset Information

Tertiary alcohol preferred: Hydroxylation of trans-3-methyl-L-proline with proline hydroxylases.

ABSTRACT: The enzymatic synthesis of tertiary alcohols by the stereospecific oxidation of tertiary alkyl centers is a most-straightforward but challenging approach, since these positions are sterically hindered. In contrast to P450-monooxygenases, there is little known about the potential of non-heme iron(II) oxygenases to catalyze such reactions. We have studied the hydroxylation of trans-3-methyl-L-proline with the ?-ketoglutarate (?-KG) dependent oxygenases, cis-3-proline hydroxylase type II and cis-4-proline hydroxylase (cis-P3H_II and cis-P4H). With cis-P3H_II, the tertiary alcohol product (3R)-3-hydroxy-3-methyl-L-proline was obtained exclusively but in reduced yield (~7%) compared to the native substrate L-proline. For cis-P4H, a complete shift in regioselectivity from C-4 to C-3 was observed so that the same product as with cis-P3H_II was obtained. Moreover, the yields were at least as good as in control reactions with L-proline (~110% relative yield). This result demonstrates a remarkable potential of non-heme iron(II) oxygenases to oxidize substrates selectively at sterically hindered positions.

SUBMITTER: Klein C

PROVIDER: S-EPMC3252868 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Tertiary alcohol preferred: Hydroxylation of trans-3-methyl-L-proline with proline hydroxylases.

Klein Christian C Hüttel Wolfgang W

Beilstein journal of organic chemistry 20111205

The enzymatic synthesis of tertiary alcohols by the stereospecific oxidation of tertiary alkyl centers is a most-straightforward but challenging approach, since these positions are sterically hindered. In contrast to P450-monooxygenases, there is little known about the potential of non-heme iron(II) oxygenases to catalyze such reactions. We have studied the hydroxylation of trans-3-methyl-L-proline with the α-ketoglutarate (α-KG) dependent oxygenases, cis-3-proline hydroxylase type II and cis-4- ...[more]

PMID: 22238542

Similar Datasets

Project description:The hydroxylation of proline residues to 4-trans-hydroxyproline (Hyp) is a common post-translational protein modification in plants, mediated by prolyl 4-hydroxylases (P4Hs). Hyps predominantly occur in a group of cell wall proteins, the Hydroxyproline-rich glycoproteins (HRGPs), where they are frequently O-glycosylated. While prolyl-hydroxylation and O-glycosylation are important, e.g. for cell wall stability, they are not desirable in plant-made pharmaceuticals. Sequence motifs recognized for prolyl-hydroxylation derived from vascular plants were proposed but did not include data from mosses, such as Physcomitrella. Here, a phylogenetic reconstruction of plant P4Hs identified six P4Hs in four subfamilies in mosses. We analysed the amino acid sequences and structural environments around Hyps in Physcomitrella utilizing 73 Hyp sites in 24 secretory proteins from multiple MS/MS datasets, and found that prolines in close proximity to other prolines, alanine, serine, threonine and valine were preferentially hydroxylated. About 95 % of the Hyp sites were predictable with a combination of previously defined motifs and methods. In our data, AOV (Ala-Hyp-Val) was the most frequent prolyl-hydroxylation pattern. Additionally, short arabinose chains were attached to Hyps in two cell-wall pectinesterases. A combination of 443 AlphaFold structure models and our MS data of peptides with nearly 3000 proline sites found Hyps predominantly on protein surfaces in disordered regions. Moss-produced human erythropoietin (EPO) exhibited plant-specific O-glycosylation with arabinose chains on two Hyps. This modification was significantly reduced in a P4H1 single knock-out (KO) Physcomitrella mutant. Quantitative proteomics after isotope labelling with different P4H-KO moss mutants revealed specific changes in the amount of proteins, including HRGPs, and a modified prolyl-hydroxylation pattern from the mutants, suggesting a differential function of the six Physcomitrella P4Hs. Quantitative RT-PCR proved a differential effect of single P4H KOs on the expression of the other five p4h genes, suggesting a partial compensation of the mutation. AlphaFold-Multimer models for Physcomitrella P4H1 and its target EPO peptide superposed with the crystal structure of Chlamydomonas P4H1 and a peptide substrate suggested significant amino acids in the active centre of the enzyme that form H-bonds with the peptide substrate, and revealed differences between P4H1 and the other five Physcomitrella P4Hs.

Dataset Information

Tertiary alcohol preferred: Hydroxylation of trans-3-methyl-L-proline with proline hydroxylases.

Publications

Tertiary alcohol preferred: Hydroxylation of trans-3-methyl-L-proline with proline hydroxylases.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets