How Similar Are P450s and What Can Their Differences Teach Us?

Graham, Sandra E.; Peterson, Julian A.

doi:10.1006/abbi.1999.1350

Cited by 185 publications

(151 citation statements)

References 38 publications

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“…As expected, HmtN adopts the typical prism-shape structure of cytochrome P450s which consists of 12 a helices and 14 b strands (Fig. 3A) [23]. The heme prosthetic group is sandwiched between I helix and cysteine-ligand loop.…”

Section: The Overall Structure Of Hmtnsupporting

confidence: 69%

Structural analysis of HmtT and HmtN involved in the tailoring steps of himastatin biosynthesis

Zhang¹,

Chen

Wang

et al. 2013

FEBS Letters

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Himastatin is a novel antibiotic featuring a bicyclohexadepsipeptide structure. On the himastatin biosynthesis pathway, three cytochrome P450s (HmtT, HmtN, HmtS) are responsible for the post‐tailoring of the cyclohexadepsipeptide backbone. Here we report the crystal structures of HmtT and HmtN. The overall structures of these two proteins are homologous to other cytochrome P450s. However, the exceptionally long F–G loop in HmtT has a highly unusual conformation and extends deep into the active site. As a result, the F/G helices of HmtT are both kinked. In contrast, the F/G helices of HmtN are straight. Also, the F/G helices in HmtT and HmtN take distinctive orientations, which may be a contributing factor for the substrate specificity of these two enzymes.

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Section: The Overall Structure Of Hmtnsupporting

confidence: 69%

Structural analysis of HmtT and HmtN involved in the tailoring steps of himastatin biosynthesis

Zhang¹,

Chen

Wang

et al. 2013

FEBS Letters

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“…The presence of two transitions could be ascribed to the presence in BMP of two sub-domains [43] while the increase of the T m values observed upon substrate addition is consistent with the additional interactions between the protein and the substrate leading to a conformational change known to decrease the flexibility of the protein [21,22].…”

Section: Differential Scanning Calorimetry (Dsc)mentioning

confidence: 54%

Subtle structural changes in the Asp251Gly/Gln307His P450 BM3 mutant responsible for new activity toward diclofenac, tolbutamide and ibuprofen

Nardo

Dell'Angelo

Catucci

et al. 2016

Archives of Biochemistry and Biophysics

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This paper reports the structure of the double mutant Asp251Gly/Gln307His (here called A2), generated by random mutagenesis, is able to produce 4'-hydroxydiclofenac, 2-hydroxyibuprofen and 4-hydroxytolbutamide from diclofenac, ibuprofen and tolbutamide, respectively. Here, we report the crystal structure of the heme domain of the mutant in the substrate-free form and in complex with the substrate N-palmitoylglycine, together with its impact on thermal stability, reduction potential and electron transfer.The substrate-free structure adopts a conformation more similar to the closed one found in the substrate-bound wild type enzyme, but with a higher degree of disorder in the region of the G-helix and F-G loop, part of the substrate access channel. This is due to the mutation Asp251Gly that breaks the salt bridge between Aps251 on Ihelix and Lys224 on G-helix, allowing the G-helix to move away from I-helix and conferring a higher degree of flexibility to this element. This subtle structural change is accompanied by long-range structural rearrangements of the active site with the rotation of Phe87 and a reorganization of catalytically important water molecules.Differential scanning calorimetry shows a population destabilized by 2.2°C compared to the wild type protein, probably reflecting the increased flexibility of part of the protein compared to WT. Moreover, a shift of the heme reduction potential by 50 mV toward positive values, a 2-folds higher first electron transfer rate in the absence of oxygen and a 4-folds higher NADPH consumption rate in the presence of oxygen as the only electron acceptors, when compared to wild type protein.The data demonstrate that a single mutation far away from the active site is able to trigger an increase in protein flexibility in a key region of the substrate access channel, shifting the conformational equilibrium toward the closed form of the protein that is ready to accept electrons and enter the P450 catalytic cycle as soon as a substrate is accepted.

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“…The structure of all enzymes resolved so far is very similar despite the low amino-acid sequence identity of < 20 % (Graham and Peterson 1999). Their tertiary structure is usually composed of 13 α-helixes and 5 β-sheets.…”

Section: P450 Monooxygenases: Whole Cell Oxidationsmentioning

confidence: 99%

Microbial P450 enzymes in biotechnology

Urlacher

Lutz-Wahl

Schmid

2004

Applied Microbiology and Biotechnology

200

100

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Oxidations are key reactions in chemical syntheses. Biooxidations using fermentation processes have already conquered some niches in industrial oxidation processes, since they allow the introduction of oxygen even into non-activated carbon atoms in a sterically and optically selective manner which is difficult or impossible to achieve by synthetic organic chemistry. Biooxidation using isolated enzymes is limited to oxidases and dehydrogenases. In addition, P450-catalyzed reactions require a constant supply of NAD(P)H which makes continuous cell-free processes very expensive. Quite recently, several groups have started to investigate cost-efficient ways which could allow the continuous supply of electrons to the heme iron. These include, for example, the use of electron mediators, direct electron supply from electrodes and enzymatic approaches. In addition, methods of protein design and directed evolution have been applied in an attempt to enhance the activity of the enzymes and improve their selectivity. The promising application of bacterial P450s as catalyzing agents in biocatalytic reactions and recent progress made in this field are covered in this review.3

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How Similar Are P450s and What Can Their Differences Teach Us?

Cited by 185 publications

References 38 publications

Structural analysis of HmtT and HmtN involved in the tailoring steps of himastatin biosynthesis

Structural analysis of HmtT and HmtN involved in the tailoring steps of himastatin biosynthesis

Subtle structural changes in the Asp251Gly/Gln307His P450 BM3 mutant responsible for new activity toward diclofenac, tolbutamide and ibuprofen

Microbial P450 enzymes in biotechnology

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