An infrared spectroscopic study of carbon monoxide bonding to ferrous cytochrome P-450

O'Keefe, David H.; Ebel, Richard E.; Peterson, Julian A.; Maxwell, John C.; Caughey, Winslow S.

doi:10.1021/bi00619a036

Cited by 77 publications

(53 citation statements)

References 58 publications

(84 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The 1969 cm ±1 band is assigned to a solvent-exposed FeCO unit (form III), where the CO ligand undergoes random interactions with the negative end of the water dipoles which is reported to broaden and blue-shift the n(CO) band [45]. In support of this assignment, a similar n(CO) band was reported for thermally denatured HRPC-CO (1964 cm ±1 ) [40], acid-denatured myoglobin-CO (1966 cm ±1 ) [46] and base-denatured cytochrome P 450 -CO (1966 cm ±1 ) [47]. A second marker of acid-denatured HRPC-CO is the reversal of the positive and negative CD bands, as seen in Fig.…”

Section: Discussionsupporting

confidence: 64%

Buffer-anion-dependent Ca2+ leaching from horseradish peroxidase at low pH

Wright¹,

English²

2001

J. Biol. Inorg. Chem.

View full text Add to dashboard Cite

Despite highly conserved active-site structures, members of the plant peroxidase superfamily exhibit a wide range of pH optima. Horseradish peroxidase isozyme C (HRPC) is an ideal peroxidase to investigate the structural determinants of pH stability and activity in superfamily members. Conflicting reports exist on the low-pH stability of HRPC and consequently the pKa of the catalytic distal histidine, which is neutral in active peroxidases. Towards resolving such discrepancies, acid-induced changes in HRPC from two popular commercial suppliers were systematically analyzed. Specifically, FTIR v(CO) and Soret-CD spectra of HRPC-CO and Soret absorption of ferric HRPC were recorded to probe time-dependent heme-pocket changes at pH 3.0 in phosphate, citrate and formate buffers, while the FTIR amide I' and far-UV CD spectra were examined to probe changes in secondary structure. Both HRPC-CO samples exhibited identical pH 7.0 v(CO) bands at 1934 and 1905 cm-1. In the pH 3.0 spectrum of sample A, the 1934 cm-1 band was dominant while a broad 1969 cm-1 band appeared in sample B. The intensity of this band, which is assigned to solvent-exposed heme, was greater in citrate than phosphate buffer, but in formate the 1934 cm-1 band remained dominant. Other spectral changes mirrored the v(CO) trends. No time- or buffer-anion-dependent conformation changes were detected in 1 mM CaCl2, revealing that buffer-anion-dependent leaching of stabilizing Ca2+ from HRPC occurs at pH 3.0. Since the N-glycans present in HRPC are of the flexible protein-surface-shielding type, the variation in low-pH conformational stability of the HRPC samples could be attributed to heterogeneous glycosylation, which was detected by SDS-PAGE. It is further proposed that glycosylation patterns may affect the low-pH stability of class II and III plant peroxidases.

show abstract

Section: Discussionsupporting

confidence: 64%

Buffer-anion-dependent Ca2+ leaching from horseradish peroxidase at low pH

Wright¹,

English²

2001

J. Biol. Inorg. Chem.

View full text Add to dashboard Cite

show abstract

“…In **. _t*~*, ..... this context it was shown that NO binds to CYP heme groups with high affinity (10) and that other hemoproteins, such as the cyclooxygenase or the lipoxygenase, are also inhibited by NO (35,36). However, in a former study we have demonstrated by electron paramagnetic resonance (EPR) that predominantly nonheme iron-nitrosyl complexes are formed by NO in hepatocytes (37).…”

Section: Resultsmentioning

confidence: 99%

“…Many of these effects are based on modulation of enzyme activity through binding of NO to prosthetic iron complexes. In this context it is interesting that NO was used for years as a spin-label probe to investigate the role of heme groups in the catalytic centers of cytochrome P450 (CYP) enzymes (10). Consequently, it was demonstrated that NO inhibits CYPdependent reactions when microsomal preparations were exposed to NO (11).…”

Section: S-nitrosylacetylpenicillamine Led To a Concentration-dependentmentioning

confidence: 99%

Inhibition of cytochromes P4501A by nitric oxide.

Stadler

Trockfeld

Schmalix

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

183

View full text Add to dashboard Cite

Inflammatory stimulation of the liver leads to the induction of nitric oxide (NO) biosynthesis. Because NO binds to the catalytic heme moiety ofcytochromes P450 (CYPs), we investigated whether NO interferes with specific CYPdependent metabolic pathways. In a first experimental approach V79 Chinese hamster cells genetically engineered for stable expression of rat and human CYPlAl and -1A2 were used. Incubation with the NO donors sodium nitroprusside and S-nitrosylacetylpenicillamine led to a concentration-dependent inhibition of all four CYP enzymes. CYPlAl was more sensitive to the inhibitory effect of NO than CYP1A2. In the second part of the study, endogenous NO synthesis was induced in rat hepatocytes by incubation with a mixture of cytokines and endotoxin. Concurrently, as NO production in hepatocytes increased within 24 hr, a decrease in CYPlAl-dependent benzo[a]pyrene turnover was observed to almost undetectable levels. The competitive inhibitor of NO synthesis, NGmonomethyl-L-arginine, was able to significantly restore CYPlAl activity in the presence of cytokines and endotoxin. Inhibition of hepatocellular CYP activity by NO was predominantly due to a direct effect on the enzymes. However, NO-dependent inhibition of CYP expression at a transcriptional level was also demonstrated. Our results indicate that inhibition of NO biosynthesis in patients suffering from systemic inflammatory response syndromes may help to restore biotransformation capacity of the liver.Hepatocellular dysfunction is a detrimental consequence of the metabolic response ofthe liver to prolonged inflammatory stimulation. Characteristic features are suppression of synthetic performance and a profound inhibition of xenobiotic biotransformation (1, 2). Therapy is not available because the molecular basis of these phenomena remains to be elucidated. However, a better understanding of the pathophysiology of the inflamed liver may result from the discovery of L-arginine-dependent nitric oxide (NO) production in hepatocytes and nonparenchymal liver cells (3, 4). As in many other cell types, an inducible NO synthase (iNOS) was identified in hepatocytes upon stimulation with cytokines and endotoxin (5, 6). Induction of iNOS was also observed in the course of parasitic infections, such as malaria (7). Recently, the gene encoding iNOS has been cloned from human hepatocytes in full length for heterologous expression (8).NO exerts a plethora of biologic functions (9). Many of these effects are based on modulation of enzyme activity through binding of NO to prosthetic iron complexes. In this context it is interesting that NO was used for years as a spin-label probe to investigate the role of heme groups in the catalytic centers of cytochrome P450 (CYP) enzymes (10). Consequently, it was demonstrated that NO inhibits CYPdependent reactions when microsomal preparations were exposed to NO (11). CYP enzymes, also referred to as microsomal monooxygenases, catalyze oxidative key reactions in the biotransformation of xenobiotics (12). Therefore, in...

show abstract

“…The P4503P420 conversion was first described by Omura and Sato (31), by using snake venom-or deoxycholate-treated rabbit liver microsomal P450 preparations. Subsequently, it has been shown that P4503P420 transitions may be induced by temperature or pressure perturbations (9,16), as well as treatment with various chemicals (17) or pH extrema (28). Some of these treatments have been exploited to investigate P450 structural architecture and prosthetic group ligand coordination (24).…”

Section: Discussionmentioning

confidence: 99%

Involvement of a Cytochrome P450 Monooxygenase in Thaxtomin A Biosynthesis by Streptomyces acidiscabies

et al. 2002

View full text Add to dashboard Cite

The biosynthesis of the thaxtomin cyclic dipeptide phytotoxins proceeds nonribosomally via the thiotemplate mechanism. Acyladenylation, thioesterification, N-methylation, and cyclization of two amino acid substrates are catalyzed by the txtAB-encoded thaxtomin synthetase. Nucleotide sequence analysis of the region 3 of txtAB in Streptomyces acidiscabies 84.104 identified an open reading frame (ORF) encoding a homolog of the P450 monooxygenase gene family. It was proposed that thaxtomin A phenylalanyl hydroxylation was catalyzed by the monooxygenase homolog. The ORF was mutated in S. acidiscabies 84.104 by using an integrative gene disruption construct, and culture filtrate extracts of the mutant were assayed for the presence of dehydroxy derivatives of thaxtomin A. Reversed-phase high-performance liquid chromatography (HPLC) and HPLC-mass spectrometry indicated that the major component in culture filtrate extracts of the mutant was less polar and smaller than thaxtomin A. Comparisons of electrospray mass spectra as well as 1 H-and 13 C-nuclear magnetic resonance spectra of the purified compound with those previously reported for thaxtomins confirmed the structure of the compound as 12,15-N-dimethylcyclo-(L-4-nitrotryptophyl-L-phenylalanyl), the didehydroxy analog of thaxtomin A. The ORF, designated txtC, was cloned and the recombinant six-His-tagged fusion protein produced in Escherichia coli and purified from cell extracts. TxtC produced in E. coli exhibited spectral properties similar to those of cytochrome P450-type hemoproteins that have undergone conversion to the catalytically inactive P420 form. Based on these properties and the high similarity of TxtC to other wellcharacterized P450 enzymes, we conclude that txtC encodes a cytochrome P450-type monooxygenase required for postcyclization hydroxylation of the cyclic dipeptide.

show abstract

An infrared spectroscopic study of carbon monoxide bonding to ferrous cytochrome P-450

Cited by 77 publications

References 58 publications

Buffer-anion-dependent Ca2+ leaching from horseradish peroxidase at low pH

Buffer-anion-dependent Ca2+ leaching from horseradish peroxidase at low pH

Inhibition of cytochromes P4501A by nitric oxide.

Involvement of a Cytochrome P450 Monooxygenase in Thaxtomin A Biosynthesis by Streptomyces acidiscabies

Contact Info

Product

Resources

About