Chemistry Reference
In-Depth Information
4 NMR in Studies of Paramagnetic Metalloproteins . . . ...................................... 84
4.1 Paramagnetism-Based Structural Restraints .......................................... 84
4.2 13 C-Detected Protonless NMR . ....................................................... 87
5 Perspectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90
1
Introduction
Metal ions play important roles in life science and the molecular mechanism of
metal-dependent life processes and all matters regarding metal within a cell or
tissue/organ are topics of the emerging fields of metallomics and metalloproteomics
[ 1 - 3 ]. Metal ions must usually associate with proteins (and other biomolecules),
i.e., metalloproteins, to prevent the toxic effects of metal excess. Metalloproteins are
one of the most diverse classes of proteins with the intrinsic metal ions providing
catalytic, regulatory, and structural roles critical to protein function, and are found in
plants, animals, and many microorganisms. It has been estimated that metalloproteins
account for approximately one-quarter to one-third of all the proteins in the human
body [ 4 ]. A recent study revealed that many of metalloproteomes still remain
uncharacterized [ 5 ]. A systematic bioinformatics survey of 1,371 metalloenzymes
with known structures showed that about 40% of enzyme-catalyzed reactions
involve metal ions, e.g., magnesium, zinc, and iron [ 6 ]. Enormous effort has been
devoted toward understanding the structure and function of metalloproteins and
such knowledge has been used to design a new functional metalloprotein [ 7 ] and to
rationalize and to search for new metalloproteins by a bioinformatics approach
[ 6 , 8 ]. However, it is impossible currently from gene sequences to predict the
numbers and types of metals an organism assimilates from its environment or
uses in its metalloproteins because the geometry and composition of metal binding
site are diverse and poorly recognized [ 9 , 10 ]. Therefore, understanding of the
function of metalloproteins comes from individual characterization of the structures
of the proteins and chemical states of the metal centers by various spectroscopic
techniques including NMR spectroscopy, circular dichroism (CD), electronic
absorption spectroscopy (UV), small angle X-ray absorption, as well as extended
X-ray absorption fine structure (EXAFS).
Over the last three decades, NMR spectroscopy has been developed into a very
important and versatile analytical technique both in the chemical and biological
sciences. It has been used within the framework of Structural Genomic (SG)
projects worldwide for determination of structures of proteins at the atomic level
under physiologically relevant conditions [ 11 - 13 ]. Moreover, NMR spectroscopy
is applicable to study of the interactions of proteins with other molecules including
proteins, nucleic acids, and even small molecules which are mainly based on the
sensitivity of the chemical shifts towards changes in chemical environments
[ 14 , 15 ]. Application of this technique to structural studies is limited to small
proteins (30-35 kDa) even with the aid of isotopic labeling ( 13 C, 15 N and 2 H)
although backbone assignments of a 723-residue enzyme with a molecular weight
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