30724-02-8Relevant academic research and scientific papers
Pneumocandin biosynthesis: Involvement of a trans-selective proline hydroxylase
Houwaart, Stefanie,Youssar, Loubna,Hüttel, Wolfgang
, p. 2365 - 2369 (2014)
Echinocandins are cyclic nonribosomal hexapeptides based mostly on nonproteinogenic amino acids and displaying strong antifungal activity. Despite previous studies on their biosynthesis by fungi, the origin of three amino acids, trans-4-and trans-3-hydroxyproline, as well as trans-3-hydroxy-4-meth-ylproline, is still unknown. Here we describe the identification, overexpression, and characterization of GloF, the first eukaryot-ic a-ketoglutarate/FeII-dependent proline hydroxylase from the pneumocandin biosynthesis cluster of the fungus Glarea loz-oyensis ATCC 74030. In in vitro transformations with L-proline, GloF generates trans-4- and trans-3-hydroxyproline simultaneously in a ratio of 8:1; the latter reaction was previously unknown for proline hydroxylase catalysis. trans-4-Methyl-L-proline is converted into the corresponding trans-3-hydroxypro-line. All three hydroxyprolines required for the biosynthesis of the echinocandins pneumocandins A0 and B0 in G. lozoyensis are thus provided by GloF. Sequence analyses revealed that GloF is not related to bacterial proline hydroxylases, and none of the putative proteins with high sequence similarity in the databases has been characterized so far.
Enzymatic production of trans-4-hydroxy-L-proline by regio- and stereospecific hydroxylation of L-proline.
Shibasaki,Mori,Ozaki
, p. 746 - 750 (2000)
A proline 4-hydroxylase gene, which was cloned from Dactylosporangium sp. RH1, was overexpressed in Escherichia coli W1485 on a plasmid under a tryptophan tandem promoter after the codon usage of the 5' end of the gene was optimized. The proline 4-hydroxylase activity was l600-fold higher than that in Dactylosporangium sp. RH1. trans-4-Hydroxy-L-proline(Hyp) was produced and accumulated to 41 g/L (87% yield from L-proline) in 100 h when the recombinant E. coli was cultivated in a medium containing L-proline and glucose. 2-Oxoglutarate, which is necessary for the hydroxylation of L-proline by proline 4-hydroxylase, was apparently supplied from glucose through the cellular metabolic pathway. The putA mutant of W1485, which is not able to degrade L-proline, has allowed the quantitative conversion of L-proline to Hyp. The formation of other isomers of hydroxyproline was not observed. Productivity of Hyp was almost the same in a larger-scale culture. The method of manufacturing Hyp from L-proline was established.
A carbohydrate based straightforward approach to trans-4-hydroxy-D-proline and trans-4-hydroxy-D-prolinol
Kumar Mishra, Umesh,Ramesh, Namakkal G.
, (2020)
Synthesis of trans-4-hydroxy-D-proline and the corresponding prolinol has been accomplished starting from 4,6-di-O-benzyl-3-deoxy-D-glucal, an enol-ether derived from D-glucose, through oxidative cleavage of a vicinal diol intermediate as the key step. This work represents a practical approach to an unnatural, yet synthetically and biologically very significant, amino acid.
Recharacterization of the mammalian cytosolic type 2 (R)-β-hydroxybutyrate dehydrogenase as 4-oxo-L-proline reductase (EC 1.1.1.104)
Bozko, Maria,Drozak, Jakub,Jagielski, Adam K.,Kocdemir, Kubra,Kwiatkowski, Sebastian,Witecka, Apolonia,Zarod, Michal
, (2022/03/23)
Early studies revealed that chicken embryos incubated with a rare analog of L-proline, 4-oxo-L-proline, showed increased levels of the metabolite 4-hydroxy-L-proline. In 1962, 4-oxo-L-proline reductase, an enzyme responsible for the reduction of 4-oxo-L-proline, was partially purified from rabbit kidneys and characterized biochemically. However, only recently was the molecular identity of this enzyme solved. Here, we report the purification from rat kidneys, identification, and biochemical characterization of 4-oxo-L-proline reductase. Following mass spectrometry analysis of the purified protein preparation, the previously annotated mammalian cytosolic type 2 (R)-βhydroxybutyrate dehydrogenase (BDH2) emerged as the only candidate for the reductase. We subsequently expressed rat and human BDH2 in Escherichia coli, then purified it, and showed that it catalyzed the reversible reduction of 4-oxo-L-proline to cis-4-hydroxy-L-proline via chromatographic and tandem mass spectrometry analysis. Specificity studies with an array of compounds carried out on both enzymes showed that 4-oxo-L-proline was the best substrate, and the human enzyme acted with 12,500-fold higher catalytic efficiency on 4-oxo-L-proline than on (R)-β-hydroxybutyrate. In addition, human embryonic kidney 293T (HEK293T) cells efficiently metabolized 4-oxo-L-proline to cis-4-hydroxy-L-proline, whereas HEK293T BDH2 KO cells were incapable of producing cis-4-hydroxy-L-proline. Both WT and KO HEK293T cells also produced trans-4-hydroxy-L-proline in the presence of 4-oxo-L-proline, suggesting that the latter compound might interfere with the trans-4-hydroxy-L-proline breakdown in human cells. We conclude that BDH2 is a mammalian 4-oxo-L-proline reductase that converts 4-oxo-L-proline to cis-4-hydroxy-L-proline and not to trans-4-hydroxy-L-proline, as originally thought. We also hypothesize that this enzyme may be a potential source of cis-4-hydroxy-L-proline in mammalian tissues.
Modular Chemoenzymatic Synthesis of GE81112 B1 and Related Analogues Enables Elucidation of Its Key Pharmacophores
Zwick, Christian R.,Sosa, Max B.,Renata, Hans
supporting information, p. 1673 - 1679 (2021/01/25)
The GE81112 complex has garnered much interest due to its broad antimicrobial properties and unique ability to inhibit bacterial translation initiation. Herein we report the use of a chemoenzymatic strategy to complete the first total synthesis of GE81112 B1. By pairing iron and α-ketoglutarate dependent hydroxylases found in GE81112 biosynthesis with traditional synthetic methodology, we were able to access the natural product in 11 steps (longest linear sequence). Following this strategy, 10 GE81112 B1 analogues were synthesized, allowing for identification of its key pharmacophores. A key feature of our medicinal chemistry effort is the incorporation of additional biocatalytic hydroxylations in modular analogue synthesis to rapidly enable exploration of relevant chemical space.
Discovery of New Fe(II)/α-Ketoglutarate-Dependent Dioxygenases for Oxidation of l-Proline
Dussauge, Solene,Moore, Charles,Snajdrova, Radka,Tassano, Erika,Vargas, Alexandra
supporting information, (2022/02/09)
Genome mining for novel Fe(II)/α-ketoglutarate-dependent dioxygenases (αKGDs) to expand the enzymatic repertoire in the oxidation of l-proline is reported. Through clustering of proteins, we predicted regio- and stereoselectivity in the hydroxylation reaction and validated this hypothesis experimentally. Two novel byproducts in the reactions with enzymes from Bacillus cereus and Streptomyces sp. were isolated, and the structures were determined to be a 3,4-epoxide and a 3,4-diol, respectively. The mechanism for the formation of the epoxide was investigated by performing an 18O-labeling experiment. We propose that the mechanism proceeds via initial cis-3-hydroxylation followed by ring closure. A biocatalytic step was run on subgram quantities of starting material without any significant optimization of the conditions. However, the substrate concentration was 40-fold higher than the usual reported titers for recombinant P450-mediated hydroxylations, showing the synthetic potential of αKGDs on a preparative scale.
Studies on the selectivity of proline hydroxylases reveal new substrates including bicycles
Smart, Tristan J.,Hamed, Refaat B.,Claridge, Timothy D.W.,Schofield, Christopher J.
supporting information, (2019/11/26)
Studies on the substrate selectivity of recombinant ferrous-iron- and 2-oxoglutarate-dependent proline hydroxylases (PHs) reveal that they can catalyse the production of dihydroxylated 5-, 6-, and 7-membered ring products, and can accept bicyclic substrates. Ring-substituted substrate analogues (such hydroxylated and fluorinated prolines) are accepted in some cases. The results highlight the considerable, as yet largely untapped, potential for amino acid hydroxylases and other 2OG oxygenases in biocatalysis.
Spiegelmeric 4R/S-hydroxy/amino-L/D-prolyl collagen peptides: conformation and morphology of self-assembled structures
Ganesh, Krishna N,More, Shahaji H
, (2020/03/11)
The primary structure of collagen, the major protein in connective tissue of mammals, comprises of repeating triads [(LPro-LHyp-Gly)n, P1, LHyp being 4R-hydroxy-lProline)] in a single strand that adopts left-handed polyproline II type helix. Three such single stranded helices wind around each another and held together by interchain H-bonds to form right-handed triple helix. This manuscript reports on collagen derived from its mirror image triad [(DPro-DHyp-Gly)n, P2, DHyp being 4S-hydroxy-DProline) and its 4-amino analogue (DPro-DAmp-Gly)n P4, DAmp being 4S-amino-DProline that form corresponding spiegelmeric triplexes. The amino L-collagen peptide (LPro-LAmp-Gly)n P3 and its D-analogue P4 show higher thermal stabilities compared to 4-hydroxy-lProline collagen peptides P1 and P2. The enantiomeric peptide pairs show mirror image CD profiles and identical thermal stability, with ionizable 4-amino group in P3 and P4 imparting pH dependent triplex stability. Upon cold mixing of the L- and D-collagen peptides, different morphological nanostructures arise from inter triplex peptide association. When the peptides are hot mixed (annealed), the inter peptide association occurs via interaction of single stranded peptide chains of opposite handedness leading to networked gel formation in P1 and P2, while the charged peptides P3 and P4 show more ordered nanofibers, different from the enantiomerically pure peptides. The nanocomposites of such chiral hybrid peptides may have not only interesting physicomorphology, but also biological properties that need exploration.
INHIBITORS OF CYCLIN DEPENDNT KINASE 7 (CDK7)
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Paragraph 339, (2018/02/28)
The present invention provides novel compounds of Formula (I) and pharmaceutically acceptable salts, solvates, hydrates, tautomers, stereoisomers, isotopically labeled derivatives, and compositions thereof. Also provided are methods and kits involving the compounds or compositions for treating or preventing proliferative diseases (e.g., cancers (e.g., leukemia, melanoma, multiple myeloma), benign neoplasms, angiogenesis, inflammatory diseases, autoinflammatory diseases, and autoimmune diseases) in a subject. Treatment of a subject with a proliferative disease using a compound or composition of the invention may inhibit the aberrant activity of cyclin-dependent kinase 7 (CDK7), and therefore, induce cellular apoptosis and/or inhibit transcription in the subject.
Discovery of new A- and B-type laxaphycins with synergistic anticancer activity
Cai, Weijing,Matthew, Susan,Chen, Qi-Yin,Paul, Valerie J.,Luesch, Hendrik
, p. 2310 - 2319 (2018/04/02)
Two new cyclic lipopeptides termed laxaphycins B4 (1) and A2 (2) were discovered from a collection of the marine cyanobacterium Hormothamnion enteromorphoides, along with the known compound laxaphycin A. The planar structures were solved based on a combined interpretation of 1D and 2D NMR data and mass spectral data. The absolute configurations of the subunits were determined by chiral LC-MS analysis of the hydrolysates, advanced Marfey's analysis and 1D and 2D ROESY experiments. Consistent with similar findings on other laxaphycin A- and B-type peptides, laxaphycin B4 (1) showed antiproliferative effects against human colon cancer HCT116 cells with IC50 of 1.7 μM, while laxaphycins A and A2 (2) exhibited weak activities. The two major compounds isolated from the sample, laxaphycins A and B4, were shown to act synergistically to inhibit the growth of HCT116 colorectal cancer cells.
