Resonance Raman studies of the stoichiometric catalytic turnover of a substrate - Stearoyl-acyl carrier protein Δ9 desaturase complex

K. S. Lyle, Pierre Moenne-Loccoz, J. Ai, J. Sanders-Loehr, T. M. Loehr, B. G. Fox

Research output: Contribution to journalArticle

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Abstract

Resonance Raman spectroscopy has been used to study the effects of substrate binding (stearoyl-acyl carrier protein, 18:0-ACP) on the diferric centers of Ricinus communis 18:0-ACP Δ9 desaturase. These studies show that complex formation produces changes in the frequencies of ν(s)(Fe-O-Fe) and ν(as)(Fe-O-Fe) consistent with a decrease in the Fe-O-Fe angle from ~ 123°in the oxo-bridged diferric centers of the as-isolated enzyme to ~ 120° in oxo-bridged diferric centers of the complex. Analysis of the shifts in ν(s)(Fe-O-Fe) and ν(as)(Fe-O-Fe) as a function of 18:0-ACP concentration also suggests that 4e--reduced Δ9D containing two diferrous centers has a higher affinity for 18:0-ACP than resting Δ9D containing two diferric centers. Catalytic turnover of a stoichiometric complex of 18:0-ACP and Δ9D was used to investigate whether an O-atom from O2 would be incorporated into a bridging position of the resultant μ-oxo-bridged diferric centers during the desaturation reaction. Upon formation of ~ 70% yield of 18:1-ACP product in the presence of 18O2, no incorporation of an 18O atom into the μ-oxo bridge position was detected. The result with 18:0-ACP Δ9 desaturase differs from that obtained during the tyrosyl radical formation reaction of the diiron enzyme ribonucleotide reductase R2 component, which proceeds with incorporation of an O-atom from O2 into the μ-oxo bridge of the resting diferric site. The possible implications of these results for the O-O bond cleavage reaction and the nature of intermediates formed during Δ9D catalysis are discussed.

Original languageEnglish (US)
Pages (from-to)10507-10513
Number of pages7
JournalBiochemistry
Volume39
Issue number34
DOIs
StatePublished - Aug 29 2000

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acyl-(acyl-carrier-protein)desaturase
Acyl Carrier Protein
Ricinus
Ribonucleotide Reductases
Atoms
Raman Spectrum Analysis
Substrates
Enzymes
Catalysis
Raman spectroscopy

ASJC Scopus subject areas

  • Biochemistry

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Resonance Raman studies of the stoichiometric catalytic turnover of a substrate - Stearoyl-acyl carrier protein Δ9 desaturase complex. / Lyle, K. S.; Moenne-Loccoz, Pierre; Ai, J.; Sanders-Loehr, J.; Loehr, T. M.; Fox, B. G.

In: Biochemistry, Vol. 39, No. 34, 29.08.2000, p. 10507-10513.

Research output: Contribution to journalArticle

Lyle, K. S. ; Moenne-Loccoz, Pierre ; Ai, J. ; Sanders-Loehr, J. ; Loehr, T. M. ; Fox, B. G. / Resonance Raman studies of the stoichiometric catalytic turnover of a substrate - Stearoyl-acyl carrier protein Δ9 desaturase complex. In: Biochemistry. 2000 ; Vol. 39, No. 34. pp. 10507-10513.
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abstract = "Resonance Raman spectroscopy has been used to study the effects of substrate binding (stearoyl-acyl carrier protein, 18:0-ACP) on the diferric centers of Ricinus communis 18:0-ACP Δ9 desaturase. These studies show that complex formation produces changes in the frequencies of ν(s)(Fe-O-Fe) and ν(as)(Fe-O-Fe) consistent with a decrease in the Fe-O-Fe angle from ~ 123°in the oxo-bridged diferric centers of the as-isolated enzyme to ~ 120° in oxo-bridged diferric centers of the complex. Analysis of the shifts in ν(s)(Fe-O-Fe) and ν(as)(Fe-O-Fe) as a function of 18:0-ACP concentration also suggests that 4e--reduced Δ9D containing two diferrous centers has a higher affinity for 18:0-ACP than resting Δ9D containing two diferric centers. Catalytic turnover of a stoichiometric complex of 18:0-ACP and Δ9D was used to investigate whether an O-atom from O2 would be incorporated into a bridging position of the resultant μ-oxo-bridged diferric centers during the desaturation reaction. Upon formation of ~ 70{\%} yield of 18:1-ACP product in the presence of 18O2, no incorporation of an 18O atom into the μ-oxo bridge position was detected. The result with 18:0-ACP Δ9 desaturase differs from that obtained during the tyrosyl radical formation reaction of the diiron enzyme ribonucleotide reductase R2 component, which proceeds with incorporation of an O-atom from O2 into the μ-oxo bridge of the resting diferric site. The possible implications of these results for the O-O bond cleavage reaction and the nature of intermediates formed during Δ9D catalysis are discussed.",
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AB - Resonance Raman spectroscopy has been used to study the effects of substrate binding (stearoyl-acyl carrier protein, 18:0-ACP) on the diferric centers of Ricinus communis 18:0-ACP Δ9 desaturase. These studies show that complex formation produces changes in the frequencies of ν(s)(Fe-O-Fe) and ν(as)(Fe-O-Fe) consistent with a decrease in the Fe-O-Fe angle from ~ 123°in the oxo-bridged diferric centers of the as-isolated enzyme to ~ 120° in oxo-bridged diferric centers of the complex. Analysis of the shifts in ν(s)(Fe-O-Fe) and ν(as)(Fe-O-Fe) as a function of 18:0-ACP concentration also suggests that 4e--reduced Δ9D containing two diferrous centers has a higher affinity for 18:0-ACP than resting Δ9D containing two diferric centers. Catalytic turnover of a stoichiometric complex of 18:0-ACP and Δ9D was used to investigate whether an O-atom from O2 would be incorporated into a bridging position of the resultant μ-oxo-bridged diferric centers during the desaturation reaction. Upon formation of ~ 70% yield of 18:1-ACP product in the presence of 18O2, no incorporation of an 18O atom into the μ-oxo bridge position was detected. The result with 18:0-ACP Δ9 desaturase differs from that obtained during the tyrosyl radical formation reaction of the diiron enzyme ribonucleotide reductase R2 component, which proceeds with incorporation of an O-atom from O2 into the μ-oxo bridge of the resting diferric site. The possible implications of these results for the O-O bond cleavage reaction and the nature of intermediates formed during Δ9D catalysis are discussed.

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