The structures ofMicrococcus lysodeikticuscatalase, its ferryl intermediate (compound II) and NADPH complex

Abstract

The crystal structure of the bacterial catalase from Micrococcus lysodeikticus has been re®ned using the gene-derived sequence both at 0.88 A Ê resolution using data recorded at 110 K and at 1.5 A Ê resolution with room-temperature data. The atomic resolution structure has been re®ned with individual anisotropic atomic thermal parameters. This has revealed the geometry of the haem and surrounding protein, including many of the H atoms, with unprecedented accuracy and has characterized functionally important hydrogen-bond interactions in the active site. The positions of the H atoms are consistent with the enzymatic mechanism previously suggested for beef liver catalase. The structure reveals that a 25 A Ê long channel leading to the haem is ®lled by partially occupied water molecules, suggesting an inherent facile access to the active site. In addition, the structures of the ferryl intermediate of the catalase, the so-called compound II, at 1.96 A Ê resolution and the catalase complex with NADPH at 1.83 A Ê resolution have been determined. Comparison of compound II and the resting state of the enzyme shows that the binding of the O atom to the iron (bond length 1.87 A Ê ) is associated with increased haem bending and is accompanied by a distal movement of the iron and the side chain of the proximal tyrosine. Finally, the structure of the NADPH complex shows that the cofactor is bound to the molecule in an equivalent position to that found in beef liver catalase, but that only the adenine part of NADPH is visible in the present structure.

X-ray absorption spectroscopic characterization of a cytochrome P450 compound II derivative

“…EXAFS studies of other compound II analogues indicated short iron-oxygen bonds in a model complex (36), in myoglobin peroxide (40), and in cytochrome c peroxidase ES complex (41), each of which has a ferryl-oxo moiety without a porphyrin radical cation. Long iron-oxygen bonds (1.84-1.92 Å) were reported from x-ray crystal structure studies of the compound II derivatives of HRP (43), myoglobin (44), and catalase (45) and also the compound I derivative of cytochrome c peroxidase (46), but Green (47) noted that the x-ray crystal structure results are in conflict with EXAFS, Raman spectral, and computational results and suggested that the long iron-oxygen bonds might have resulted from reduction of the ferryl species to ferric or ferrous species in the x-ray beam.…”

Crystallographic and Spectroscopic Studies of Peroxide-derived Myoglobin Compound II and Occurrence of Protonated FeIV–O

“…Refinements, including rigid body, simulated annealing, and conjugate gradient minimizations with CNS, and model improvements in O (63), were performed initially for some structures. Further steps included multiple cycles of restrained refinement in Refmac (58,64) and model building in Coot (65) and addition of water molecules by ARP/ wARP (66). In the later stages TLS refinement was introduced, and finally restrained anisotropic refinement performed with Refmac (67,68).…”

Structure, Protonation State and Dynamics of Catalase Compound II

“…[4] As both intermediates feature a Fe IV =O bond, it is expected that they have a similar FeÀO bond length. However, two recent high-resolution crystal structures of Cpds I and II give quite different FeÀO distances, 1.67 and 1.86 , respectively, [4,6,7] with a r.m.s.d. of 0.02 .…”

“…Figure 1. Structure around the active site of heme catalases (the residue numbering of MLC is used [6] ). Scheme 1.…”