Metalloproteomics

Metalloproteins are one of the most diverse classes of proteins that contain metal atoms. The function of these metalloproteins critically depends on the specific interactions between the proteins and the binding metals, such as Cu, Fe, Zn, or Mo. The intrinsic metal atoms in protein structures provide catalytic, regulatory, and structural roles critical to protein function, ranging from electron transfer, substrate binding and activation, transport and storage processes, to regulation of enzymatic activity and gene expression. 

In addition, intrinsic transition metal atoms can be used for crystallographic phasing in favorable cases based on the anomalous signal from the metal atom. Searching the Protein Data Bank (PDB) shows that almost one-fourth of the 

As mentioned above, Zn is the most abundant transition metal in cells, and plays a vital role in the functionalities of more than 300 enzymes, in the stabilization of DNA, and in gene expression. Other trace metals, such as Se, W, and Mo, are essential in human health and also important in the environment. For example, Se is usually incorporated into antioxidant enzymes as seleno-cysteine (SeCys), the redox active site of SeCys-containing enzymes, and plays a key role in host oxidative defense. It is predicted that there are a total of 25 SeCys-containing proteins in humans. The most widely known SeCys-containing enzymes are thioredoxin reductase and glutathione peroxidase. Both are ubiquitous and found in bacteria, plants, and mammals, including humans.  

When work as the biological catalysts to regulate the biological reactions and physiological functions in biological cells and organs, metalloproteins are called metalloenzymes (lUPAC definition An enzyme that, in the active state, contains one or more metal ions which are essential for its biological function.). Some typical metalloenzymes and metalloproteins are summarized in Table 1.2. 

As shown in Table 1.2, metalloenzymes contain specific numbers of metal ions at the active sites in specific proteins. The presence of metal ions allows metalloenzymes to perform functions as biocatalysts for specific enzymatic reactions including gene (DNA, RNA) synthesis, metabolism, antioxidation and so forth. 

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Szpunar J. Advances in analytical methodology for bioinorganic speciation analysis metallomics, metalloproteomics and heteroatom-tagged proteomics and metabolomics. Analyst 2005 130 442-465. 

Andreini C, Bertini I, Rosato A (2009) Metalloproteomes a bioinformatic approach. Acc ChemRes 42 1471-1479... 

Metallomics and metalloproteomics are emerging fields addressing the role, uptake, transport, and storage of trace metals essential for protein functions. The methodologies utilized in... 

Compared to the well known genomics and proteomics, metallomics and metalloproteomics are relatively new fields. They are receiving great attention in the investigation of trace elements in biology and expected to develop as an interdisciplinary science complementary to genomics and proteomics. In the... 

Metalloproteomics is a new subject focusing on the distributions and compositions of all metalloproteins in a proteome (metalloproteome), their structural and functional characterization and their structural metal binding moieties. The specificity of metalloproteomics studies demands the need for a description of the metal-binding sites, metal stoichiometry, and metal-depen-dent structure or conformation changes as well as the identifieation and quantification of the metalloproteins. The metalloproteome can be considered as not only a subset of the metallome, but also a very important subset of the proteome. 

Figure 1.7 Roadmap for metalloproteomics studies used in the Laboratory of Nuclear Analytical Techniques of Institute of High Energy Physics, China. 2007 The Royal Society of Chemistry.

Nuclear Analytical Techniques for Metallome and Metalloproteome Distribution... 

Isotopic techniques will continuously play an important role in the emerging field of metallomics or metalloproteomics. In future, the application of isotopic tracers will probably increase and contribute significantly to the quest for new knowledge. Nevertheless, radioactive experiments need well-trained professionals in a strictly protective environment. In addition, the isotopic effect and radiation effect have to be considered. For more details about this method and the application of these techniques, readers are encouraged to read Chapter 4 (Isotopic Techniques Combined with ICP-MS and ESI-MS) in this book. 

Nuclear Analytical Techniques for the Structural Analysis of Metallomes and Metalloproteomes... 

Techniques such as ray-based techniques and nuclear magnetic resonance (NMR) can be used for the structural characterization of metallomes and metalloproteome. Ray-based techniques can characterize the structure at the atomic level. Rays that can be used for structural analysis include X-rays, gamma rays, or neutron beams. 

In the X-ray-based techniques. X-ray crystallography is the most powerful tool for the determination of macromolecular 3D structures at a resolution of 0.15-2 nm but the requirement for a single crystal will greatly limit its application to numerous biological samples. The application of X-ray crystallography for metalloproteomics is illustrated in Chapter 7 (Protein Crystallography for Metalloproteins) in this book. 

X-ray absorption spectroscopy (XAS), especially extended X-ray absorption fine structure (EXAFS), may provide an alternative tool for determining the local structure around certain atoms at a resolution of 10 to 10 nm without the requirement for crystalline samples. For example, Hg in human hair and blood samples from long-term mercury-exposed populations has been studied using EXAFS and structural information such as bond distances and coordination numbers of Hg were obtained. Further, EXAFS can provide a refinement of the structure determined from X-ray crystallography since EXAFS has higher spatial resolution than X-ray crystallography especially in local structures. More detailed information about XAS and its application in metallomics and metalloproteomics study can be found in Chapter 6 (X-ray Absorption Spectroscopy) in this book. 

Figure 1.10 Schematic flowchart for the two alternate workflows for high-throughput X-ray absorption spectroscopy (HT-XAS). Synchrotron radiation analysis consists of mapping the metal distribution using X-ray fluorescence (XRF), using XANES for metal speciation and using EXAFS for metal-site structural analysis of the metalloproteome. 2005 International Union of Crystallography.

Table 1.5 Features of the main nuclear analytical techniques for chemical element imaging, quantification, and speciation in metallmoics and metalloproteomics studies. 

This book is written by experts from disciplines as diverse as analytical chemistry, nuclear chemistry, environmental science, molecular biology, and medicinal chemistry in order to identify potential hot spots of metallomics and metalloproteomics. The scientific fundamentals of new approaches, like isotopic techniques combined with ICP-MS/ESI-MS/MS, the synchrotron radiation-based techniques. X-ray absorption spectroscopy, X-ray diffraction, and neutron scattering, as well as their various applications, with a focus on mercury, selenium, chromium, arsenic, iron and metal-based medicines are critically reviewed, which can help to understand their impacts on human health. The book will be of particular interest to researchers in the fields of environmental and industrial chemistry, biochemistry, nutrition, toxicology, and medicine. Basically, the book has two aims. The first deals with the educational point of view. Chapters 2 to 7 provide the basic concept of each of the selected nuclear analytical techniques and should be understandable by Master and PhD students in chemistry, physics, biology and nanotechnology. The... 

X-ray fluorescence is a non-destructive and multielemental analytical technique. Because of its excellent analytical sensitivity and spatial resolution under micro-beam conditions, the technique is capable of microscopic analysis, supplying information about two-dimensional (2D) distributions of trace elements. The technique can, thus, be used for imaging trace elements in biological specimens, and for the direct determination of trace elements in protein bands after slab-gel electrophoresis (GE), which is the benchmark for high-resolution protein separation, particularly in 2D format. Therefore, XRF is a useful technique for metallomics and metalloproteomics studies. 

Several variants of EDXRF with different performance and character are developed by changing the beam size of primary X-rays or the geometrical setup of the XRF instrument. Two of them, micro-XRF and total reflection XRF, will be introduced for their wide use or attractive prospects in metallo-mics and metalloproteomics fields. 

Figure 4.3 The hyphenated techniques for metallomics and metalloproteomics 2005 Wiley Periodicals, Inc.

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