Dataset Information

Functional expression and purification of DoxA, a key cytochrome P450 from Streptomyces peucetius ATCC 27952.

ABSTRACT: The antitumor drug doxorubicin is widely used in clinical practice. However, the low yield and high cost of this drug highlight the urgent need for cost-effective processes to rapidly manufacture antitumor drugs at scale. In the biosynthesis pathway, the multi-functional cytochrome P450 enzyme DoxA catalyzes the last three steps of hydroxylation. The final conversion of daunorubicin to doxorubicin is the rate-limiting step. In our work, the DoxA has been expressed with the ferredoxin reductase FDR2 and the ferredoxin FDX1 and purified to homogeneous. The reduced carbon monoxide difference spectroscopy, heme concentration, and enzymatic characteristic were characterized. These studies suggest an approach for engineering Streptomyces P450s with functional expression for mechanistic and structural studies.

SUBMITTER: Yang L

PROVIDER: S-EPMC9675340 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Functional expression and purification of DoxA, a key cytochrome P450 from <i>Streptomyces peucetius</i> ATCC 27952.

Yang Liyan L Yang Dengfeng D Wang Qingyan Q Li Juan J Li Hong-Liang HL Pan Lixia L

PeerJ 20221116

The antitumor drug doxorubicin is widely used in clinical practice. However, the low yield and high cost of this drug highlight the urgent need for cost-effective processes to rapidly manufacture antitumor drugs at scale. In the biosynthesis pathway, the multi-functional cytochrome P450 enzyme DoxA catalyzes the last three steps of hydroxylation. The final conversion of daunorubicin to doxorubicin is the rate-limiting step. In our work, the DoxA has been expressed with the ferredoxin reductase F ...[more]

PMID: 36411834

Similar Datasets

Project description:BackgroundAs well known, both natural and synthetic steroidal compounds are powerful endocrine disrupting compounds (EDCs) which can cause reproductive toxicity and affect cellular development in mammals and thus are generally regarded as serious contributors to water pollution. Streptomyces virginiae IBL14 is an effective degradative strain for many steroidal compounds and can also catalyze the C25 hydroxylation of diosgenin, the first-ever biotransformation found on the F-ring of diosgenin.ResultsTo completely elucidate the hydroxylation function of cytochrome P450 genes (CYPs) found during biotransformation of steroids by S. virginiae IBL14, the whole genome sequencing of this strain was carried out via 454 Sequencing Systems. The analytical results of BLASTP showed that the strain IBL14 contains 33 CYPs, 7 ferredoxins and 3 ferredoxin reductases in its 8.0 Mb linear chromosome. CYPs from S. virginiae IBL14 are phylogenetically closed to those of Streptomyces sp. Mg1 and Streptomyces sp. C. One new subfamily was found as per the fact that the CYP Svu001 in S. virginiae IBL14 shares 66% identity only to that (ZP_05001937, protein identifer) from Streptomyces sp. Mg1. Further analysis showed that among all of the 33 CYPs in S. virginiae IBL14, three CYPs are clustered with ferredoxins, one with ferredoxin and ferredoxin reductase and three CYPs with ATP/GTP binding proteins, four CYPs arranged with transcriptional regulatory genes and one CYP located on the upstream of an ATP-binding protein and transcriptional regulators as well as four CYPs associated with other functional genes involved in secondary metabolism and degradation.ConclusionsThese characteristics found in CYPs from S. virginiae IBL14 show that the EXXR motif in the K-helix is not absolutely conserved in CYP157 family and I-helix not absolutely essential for the CYP structure, too. Experimental results showed that both CYP Svh01 and CYP Svu022 are two hydroxylases, capable of bioconverting diosgenone into isonuatigenone and β-estradiol into estriol, respectively.

Dataset Information

Functional expression and purification of DoxA, a key cytochrome P450 from Streptomyces peucetius ATCC 27952.

Publications

Functional expression and purification of DoxA, a key cytochrome P450 from <i>Streptomyces peucetius</i> ATCC 27952.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets