Applications of X-ray absorption spectroscopy and low temperature XMCD to metalloproteins

Technical Report · Mon Jan 01 04:00:00 EST 1996

DOI:https://doi.org/10.2172/266644· OSTI ID:266644

Christiansen, J H ^[1]

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1. Univ. of California, Davis, CA (United States). Dept. of Applied Science

The author has used the extended X-ray absorption fine structure (EXAFS) and ultra-low temperature X-ray magnetic circular dichroism (XMCD) to study the environments of the metal sites in metalloproteins. EXAFS has been used to study the Zn site in spinach carbonic anhydrase. The EXAFS, in parallel with site directed mutagenesis studies, indicate that the active site Zn is in a cys-cys-his-H{sub 2}O environment, very different from the mammalian carbonic anhydrase active site. Nitrogenase, the primary enzyme in biological nitrogen fixation, contains two complex metal clusters of unique structure. EXAFS studies at the Fe and Mo K-edges of nitrogenase solutions and crystals yielded information about the various metal-metal distances in these two clusters. The author assigned 4 Fe and 3 Mo interactions >4 {angstrom}. Single crystal Mo K-edge EXAFS then found a very long Fe-Fe distance of {approximately}5.1 {angstrom}. These distances were then used to further refine the proposed crystallographic models to their highest accuracy yet. Studies were carried further by examining nitrogenas in oxidized and reduced forms--states for which there is no crystallographic information. Small structural changes were observed and an EXAFS model was put forth that attempts to deconvolute the EXAFS distances of the two metal clusters. Nitrogenase Apo I, a genetic mutant of nitrogenase which is though to contain only one of the two different metal clusters, was also examined using EXAFS. These studies showed results consistent with current models, yet the metal clusters were very disordered. Finally, ultra-low temperature methods were used to further the development of XMCD as a technique for studying biological systems. Experiments were performed on the copper in plastocyanin. Data was collected that definitively proves that the sample surface was at 0.55 {+-} 0.05 K. This result opens the door to further study of more complex biological metal clusters.

• Research Organization: Lawrence Berkeley Lab., CA (United States)
• Sponsoring Organization: USDOE, Washington, DC (United States); National Insts. of Health, Bethesda, MD (United States); National Science Foundation, Washington, DC (United States); Department of Agriculture, Washington, DC (United States)
• DOE Contract Number: AC03-76SF00098
• OSTI ID: 266644
• Report Number(s): LBL--38410; ON: DE96013149; CNN: NIH Contract GM-44380; NSF Contracts DMB-9107312; DIR-9105323; DIR-9317942; DOA Grant DOA-91-37305-6514
• Country of Publication: United States
• Language: English

Related Subjects

40 CHEMISTRY
55 BIOLOGY AND MEDICINE
BASIC STUDIES
ABSORPTION SPECTROSCOPY
COPPER COMPOUNDS
CRYSTAL STRUCTURE
EXPERIMENTAL DATA
IRON COMPOUNDS
MAGNETIC CIRCULAR DICHROISM
METALLOPROTEINS
MOLYBDENUM COMPOUNDS
NITROGENASE
STRUCTURAL CHEMICAL ANALYSIS
X-RAY SPECTROMETERS
X-RAY SPECTROSCOPY
ZINC COMPOUNDS