Abstract
Vibrational spectroscopy and in particular, resonance Raman (RR) spectroscopy, can provide molecular details on metalloproteins containing multiple cofactors, which are often challenging for other spectroscopies. Due to distinct spectroscopic fingerprints, RR spectroscopy has a unique capacity to monitor simultaneously and independently different metal cofactors that can have particular roles in metalloproteins. These include e.g., (i) different types of hemes, for instance hemes c, a and a3 in caa3-type oxygen reductases, (ii) distinct spin populations, such as electron transfer (ET) low-spin (LS) and catalytic high-spin (HS) hemes in nitrite reductases, (iii) different types of Fe-S clusters, such as 3Fe-4S and 4Fe-4S centers in di-cluster ferredoxins, and (iv) bi-metallic center and ET Fe-S clusters in hydrogenases. IR spectroscopy can provide unmatched molecular details on specific enzymes like hydrogenases that possess catalytic centers coordinated by CO and CN− ligands, which exhibit spectrally well separated IR bands. This article reviews the work on metalloproteins for which vibrational spectroscopy has ensured advances in understanding structural and mechanistic properties, including multiple heme-containing proteins, such as nitrite reductases that house a notable total of 28 hemes in a functional unit, respiratory chain complexes, and hydrogenases that carry out the most fundamental functions in cells.
Highlights
Vibrational spectroscopy has, over the past couple of decades, provided valuable information on structure-function relationship in proteins, contributing to the elucidation of molecular mechanisms, dynamics and interactions on the level that goes beyond high-resolution crystallographic structures
It does not depend on size or paramagnetic properties of the protein like nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopies, and it can be coupled with electrochemical methods and employed in time-resolved mode, offering the possibility to probe redox and transient molecular events down to the femtosecond time scale [1]
resonance Raman (RR) spectra of Fe-S cluster-containing proteins, obtained with a laser wavelength that matches the energy of S → Fe charge transfer (CT) transitions, show selectively enhanced modes involving the metal–ligand stretching coordinates in the low-frequency (200–450 cm−1) region (Table 1)
Summary
Vibrational spectroscopy has, over the past couple of decades, provided valuable information on structure-function relationship in proteins, contributing to the elucidation of molecular mechanisms, dynamics and interactions on the level that goes beyond high-resolution crystallographic structures. RR spectra of Fe-S cluster-containing proteins, obtained with a laser wavelength that matches the energy of S → Fe charge transfer (CT) transitions, show selectively enhanced modes involving the metal–ligand stretching coordinates in the low-frequency (200–450 cm−1) region (Table 1). We (i) discuss multiple hemecontaining proteins, such as nitrite reductases harboring an impressive total of 28 hemes in a functional unit, (ii) describe the work on Complex III and dihemic CcPs, the heme groups of which can be distinguished via selective reduction due to large differences in the respective redox potentials, and (iii) dwell on oxygen reductases for which RR and SERR spectroscopy played a crucial role in disentangling the structure and formation kinetics of catalytic intermediates. The stretching coordinates of the 6cLS-NO d1 heme have been observed at unusually high frequencies (585 cm−1 in Pseudomonas aeruginosa cd1NiR) in comparison to other heme–NO complexes [29,30], which is thought to be related with the electronic properties and highly ruffled structure of heme d1 [27]
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