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1.
Chemistry ; 29(42): e202301869, 2023 Jul 26.
Article in English | MEDLINE | ID: mdl-37350118

ABSTRACT

Invited for the cover of this issue are the groups of Gonzalo Jiménez-Osés and Fernando López-Gallego at CIC bioGUNE and CIC biomaGUNE, respectively. The image depicts the substrate scope of an engineered acyl transferases for the synthesis of statin derivatives. Read the full text of the article at 10.1002/chem.202300911.


Subject(s)
Hydroxymethylglutaryl-CoA Reductase Inhibitors , Acyltransferases
2.
Chemistry ; 29(42): e202300911, 2023 Jul 26.
Article in English | MEDLINE | ID: mdl-37139626

ABSTRACT

This study identifies new acyl donors for manufacturing statin analogues through the acylation of monacolin J acid by the laboratory evolved acyltransferase LovD9. Vinyl and p-nitrophenyl esters have emerged as alternate substrates for LovD9-catalyzed acylation. While vinyl esters can reach product yields as high as the ones obtained by α-dimethyl butyryl-S-methyl-3-mercaptopropionate (DMB-SMMP), the thioester for which LovD9 was evolved, p-nitrophenyl esters display a reactivity even higher than DMB-SMMP for the first acylation step yet the acylation product yield is lower. The reaction mechanisms were elucidated through quantum mechanics (QM) calculations.


Subject(s)
Hydroxymethylglutaryl-CoA Reductase Inhibitors , Acyltransferases/metabolism , Biocatalysis , Acylation , Esters , Substrate Specificity
3.
Angew Chem Int Ed Engl ; 62(13): e202218312, 2023 03 20.
Article in English | MEDLINE | ID: mdl-36718873

ABSTRACT

In vitro biosynthetic pathways that condense and reduce molecules through coenzyme A (CoASH) activation demand energy and redox power in the form of ATP and NAD(P)H, respectively. These coenzymes must be orthogonally recycled by ancillary reactions that consume chemicals, electricity, or light, impacting the atom economy and/or the energy consumption of the biosystem. In this work, we have exploited vinyl esters as dual acyl and electron donor substrates to synthesize ß-hydroxy acids through a non-decarboxylating Claisen condensation, reduction and hydrolysis stepwise cascade, including a NADH recycling step, catalyzed by a total of 4 enzymes. Herein, the chemical energy to activate the acyl group with CoASH and the redox power for the reduction are embedded into the vinyl esters. Upon optimization, this self-sustaining cascade reached a titer of (S)-3-hydroxy butyrate of 24 mM without requiring ATP and simultaneously recycling CoASH and NADH. This work illustrates the potential of in vitro biocatalysis to transform simple molecules into multi-functional ones.


Subject(s)
Hydroxy Acids , NAD , NAD/metabolism , Esters , Coenzyme A/metabolism , Adenosine Triphosphate/metabolism
4.
Sci Rep ; 7: 44834, 2017 03 23.
Article in English | MEDLINE | ID: mdl-28333133

ABSTRACT

Nucleoside reverse transcriptase (RT) inhibitors constitute the backbone of current therapies against human immunodeficiency virus type 1 and type 2 (HIV-1 and HIV-2, respectively). However, mutational pathways leading to the development of nucleoside analogue resistance are different in both types of HIV. In HIV-2, resistance to all approved nucleoside analogues is conferred by the combination of RT substitutions K65R, Q151M and M184V. Nucleotide incorporation kinetic analyses of mutant and wild-type (WT) HIV-2 RTs show that the triple-mutant has decreased catalytic efficiency due to the presence of M184V. Although similar effects were previously reported for equivalent mutations in HIV-1 RT, the HIV-2 enzymes were catalytically less efficient. Interestingly, in highly divergent HIV-1 RTs, K65R confers several-fold increased accuracy of DNA synthesis. We have determined the intrinsic fidelity of DNA synthesis of WT HIV-2 RT and mutants K65R and K65R/Q151M/M184V. Our results show that those changes in HIV-2 RT have a relatively small impact on nucleotide selectivity. Furthermore, we found that there were less than two-fold differences in error rates obtained with forward mutation assays using mutant and WT HIV-2 RTs. A different conformation of the ß3-ß4 hairpin loop in HIV-1 and HIV-2 RTs could probably explain the differential effects of K65R.


Subject(s)
HIV Infections/virology , HIV Reverse Transcriptase/genetics , HIV-1/drug effects , HIV-1/genetics , HIV-2/drug effects , HIV-2/genetics , Mutation , Alleles , Amino Acid Substitution , Anti-HIV Agents/pharmacology , Codon , DNA Replication , Drug Resistance, Viral , HIV Reverse Transcriptase/metabolism , Humans , Kinetics , Reverse Transcriptase Inhibitors/pharmacology , Reverse Transcription
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