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Chemical Reviews Apr 2014
Review
Topics: Electron Transport; Electrons; Kinetics; Metalloproteins; Models, Molecular; Protein Structure, Tertiary; Thermodynamics
PubMed: 24279515
DOI: 10.1021/cr4004715 -
Accounts of Chemical Research May 2021Rigorous substrate selectivity is a hallmark of enzyme catalysis. This selectivity is generally ascribed to a thermodynamically favorable process of substrate binding to... (Review)
Review
Rigorous substrate selectivity is a hallmark of enzyme catalysis. This selectivity is generally ascribed to a thermodynamically favorable process of substrate binding to the enzyme active site based upon complementary physiochemical characteristics, which allows both acquisition and orientation. However, this chemical selectivity is more difficult to rationalize for diminutive molecules that possess too narrow a range of physical characteristics to allow either precise positioning or discrimination between a substrate and an inhibitor. Foremost among these small molecules are dissolved gases such as H, N, O, CO, CO, NO, NO, NH, and CH so often encountered in metalloenzyme catalysis. Nevertheless, metalloenzymes have evolved to metabolize these small-molecule substrates with high selectivity and efficiency.The soluble methane monooxygenase enzyme (sMMO) acts upon two of these small molecules, O and CH, to generate methanol as part of the C1 metabolic pathway of methanotrophic organisms. sMMO is capable of oxidizing many alternative hydrocarbon substrates. Remarkably, however, it will preferentially oxidize methane, the substrate with the fewest discriminating physical characteristics and the strongest C-H bond. Early studies led us to broadly attribute this specificity to the formation of a "molecular sieve" in which a methane- and oxygen-sized tunnel provides a size-selective route from bulk solvent to the completely buried sMMO active site. Indeed, recent cryogenic and serial femtosecond ambient temperature crystallographic studies have revealed such a route in sMMO. A detailed study of the sMMO tunnel considered here in the context of small-molecule tunnels identified in other metalloenzymes reveals three discrete characteristics that contribute to substrate selectivity and positioning beyond that which can be provided by the active site itself. Moreover, the dynamic nature of many tunnels allows an exquisite coordination of substrate binding and reaction phases of the catalytic cycle. Here we differentiate between the highly selective molecular tunnel, which allows only the one-dimensional transit of small molecules, and the larger, less-selective channels found in typical enzymes. Methods are described to identify and characterize tunnels as well as to differentiate them from channels. In metalloenzymes which metabolize dissolved gases, we posit that the contribution of tunnels is so great that they should be considered to be extensions of the active site itself. A full understanding of catalysis by these enzymes requires an appreciation of the roles played by tunnels. Such an understanding will also facilitate the use of the enzymes or their synthetic mimics in industrial or pharmaceutical applications.
Topics: Biocatalysis; Catalytic Domain; Metalloproteins; Models, Molecular; Oxygenases; Small Molecule Libraries
PubMed: 33886257
DOI: 10.1021/acs.accounts.1c00058 -
Journal of Inorganic Biochemistry Sep 2022Metalloproteins represent a substantial fraction of the proteome where they have an outsized contribution to enzymology. This stems from the reactivity of transition... (Review)
Review
Metalloproteins represent a substantial fraction of the proteome where they have an outsized contribution to enzymology. This stems from the reactivity of transition metals found in the active sites of numerous classes of enzymes that undergo redox and/or spin-state transitions. Notwithstanding, NMR structures of metalloproteins deposited in the PDB are under-represented and NMR studies exploring paramagnetic states are a minute fraction of the overall database content. This state of affairs contrasts with the early recognition that paramagnetic proteins offer unique opportunities for structure-function studies which are not available for diamagnetic proteins. Recent development of novel pulse sequences that minimize quenching of signal intensity that arises from the presence of a paramagnetic center in metalloproteins is extending even further the range of systems which can be studied by solution-state NMR. In this manuscript we review solution-state NMR applications to paramagnetic proteins, highlighting the developments in both methodologies and data interpretation, laying bare the vast range of opportunities for paramagnetic NMR to contribute to the understanding of structure and function of metalloenzymes and biomimetic metallocatalysts.
Topics: Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Metalloproteins
PubMed: 35636014
DOI: 10.1016/j.jinorgbio.2022.111871 -
Nature Chemistry Mar 2018Random mutagenesis has the potential to optimize the efficiency and selectivity of protein catalysts without requiring detailed knowledge of protein structure; however,...
Random mutagenesis has the potential to optimize the efficiency and selectivity of protein catalysts without requiring detailed knowledge of protein structure; however, introducing synthetic metal cofactors complicates the expression and screening of enzyme libraries, and activity arising from free cofactor must be eliminated. Here we report an efficient platform to create and screen libraries of artificial metalloenzymes (ArMs) via random mutagenesis, which we use to evolve highly selective dirhodium cyclopropanases. Error-prone PCR and combinatorial codon mutagenesis enabled multiplexed analysis of random mutations, including at sites distal to the putative ArM active site that are difficult to identify using targeted mutagenesis approaches. Variants that exhibited significantly improved selectivity for each of the cyclopropane product enantiomers were identified, and higher activity than previously reported ArM cyclopropanases obtained via targeted mutagenesis was also observed. This improved selectivity carried over to other dirhodium-catalysed transformations, including N-H, S-H and Si-H insertion, demonstrating that ArMs evolved for one reaction can serve as starting points to evolve catalysts for others.
Topics: Biocatalysis; Metalloproteins; Molecular Structure; Mutagenesis; Polymerase Chain Reaction
PubMed: 29461523
DOI: 10.1038/nchem.2927 -
Current Opinion in Chemical Biology Feb 2011About one-third of all proteins are associated with a metal. Metalloproteomics is defined as the structural and functional characterization of metalloproteins on a... (Review)
Review
About one-third of all proteins are associated with a metal. Metalloproteomics is defined as the structural and functional characterization of metalloproteins on a genome-wide scale. The methodologies utilized in metalloproteomics, including both forward (bottom-up) and reverse (top-down) technologies, to provide information on the identity, quantity, and function of metalloproteins are discussed. Important techniques frequently employed in metalloproteomics include classical proteomic tools such as mass spectrometry and 2D gels, immobilized-metal affinity chromatography, bioinformatic sequence analysis and homology modeling, X-ray absorption spectroscopy and other synchrotron radiation based tools. Combinative applications of these techniques provide a powerful approach to understand the function of metalloproteins.
Topics: Animals; Computational Biology; Humans; Metalloproteins; Protein Conformation; Proteomics
PubMed: 21130021
DOI: 10.1016/j.cbpa.2010.11.004 -
Metallomics : Integrated Biometal... Oct 2012How cells ensure correct metallation of a given protein and whether a degree of promiscuity in metal binding has evolved are largely unanswered questions. In a classic... (Review)
Review
How cells ensure correct metallation of a given protein and whether a degree of promiscuity in metal binding has evolved are largely unanswered questions. In a classic case, iron- and manganese-dependent superoxide dismutases (SODs) catalyze the disproportionation of superoxide using highly similar protein scaffolds and nearly identical active sites. However, most of these enzymes are active with only one metal, although both metals can bind in vitro and in vivo. Iron(ii) and manganese(ii) bind weakly to most proteins and possess similar coordination preferences. Their distinct redox properties suggest that they are unlikely to be interchangeable in biological systems except when they function in Lewis acid catalytic roles, yet recent work suggests this is not always the case. This review summarizes the diversity of ways in which iron and manganese are substituted in similar or identical protein frameworks. As models, we discuss (1) enzymes, such as epimerases, thought to use Fe(II) as a Lewis acid under normal growth conditions but which switch to Mn(II) under oxidative stress; (2) extradiol dioxygenases, which have been found to use both Fe(II) and Mn(II), the redox role of which in catalysis remains to be elucidated; (3) SODs, which use redox chemistry and are generally metal-specific; and (4) the class I ribonucleotide reductases (RNRs), which have evolved unique biosynthetic pathways to control metallation. The primary focus is the class Ib RNRs, which can catalyze formation of a stable radical on a tyrosine residue in their β2 subunits using either a di-iron or a recently characterized dimanganese cofactor. The physiological roles of enzymes that can switch between iron and manganese cofactors are discussed, as are insights obtained from the studies of many groups regarding iron and manganese homeostasis and the divergent and convergent strategies organisms use for control of protein metallation. We propose that, in many of the systems discussed, "discrimination" between metals is not performed by the protein itself, but it is instead determined by the environment in which the protein is expressed.
Topics: Bacterial Proteins; Binding Sites; Iron; Iron-Sulfur Proteins; Manganese; Metalloproteins; Models, Molecular; Nonheme Iron Proteins; Racemases and Epimerases; Ribonucleotide Reductases
PubMed: 22991063
DOI: 10.1039/c2mt20142a -
Journal of Inorganic Biochemistry Apr 2014Mycobacterium tuberculosis, the pathogen that causes tuberculosis, has evolved sophisticated mechanisms for evading assault by the human host. This review focuses on M.... (Review)
Review
Mycobacterium tuberculosis, the pathogen that causes tuberculosis, has evolved sophisticated mechanisms for evading assault by the human host. This review focuses on M. tuberculosis regulatory metalloproteins that are sensitive to exogenous stresses attributed to changes in the levels of gaseous molecules (i.e., molecular oxygen, carbon monoxide and nitric oxide) to elicit an intracellular response. In particular, we highlight recent developments on the subfamily of Whi proteins, redox sensing WhiB-like proteins that contain iron-sulfur clusters, sigma factors and their cognate anti-sigma factors of which some are zinc-regulated, and the dormancy survival regulon DosS/DosT-DosR heme sensory system. Mounting experimental evidence suggests that these systems contribute to a highly complex and interrelated regulatory network that controls M. tuberculosis biology. This review concludes with a discussion of strategies that M. tuberculosis has developed to maintain redox homeostasis, including mechanisms to regulate endogenous nitric oxide and carbon monoxide levels.
Topics: Carbon Monoxide; Gene Expression Regulation, Bacterial; Heme; Humans; Hypoxia; Metalloproteins; Mycobacterium tuberculosis; Nitric Oxide; Oxidation-Reduction; Oxygen; Tuberculosis
PubMed: 24314844
DOI: 10.1016/j.jinorgbio.2013.11.003 -
International Journal of Molecular... Feb 2022Cadmium (Cd) pollution in cultivated land is caused by irresistible geological factors and human activities; intense diffusion and migration have seriously affected the... (Review)
Review
Cadmium (Cd) pollution in cultivated land is caused by irresistible geological factors and human activities; intense diffusion and migration have seriously affected the safety of food crops. Plants have evolved mechanisms to control excessive influx of Cd in the environment, such as directional transport, chelation and detoxification. This is done by some specific metalloproteins, whose key amino acid motifs have been investigated by scientists one by one. The application of powerful cell biology, crystal structure science, and molecular probe targeted labeling technology has identified a series of protein families involved in the influx, transport and detoxification of the heavy metal Cd. This review summarizes them as influx proteins (NRAMP, ZIP), chelating proteins (MT, PDF), vacuolar proteins (CAX, ABCC, MTP), long-distance transport proteins (OPT, HMA) and efflux proteins (PCR, ABCG). We selected representative proteins from each family, and compared their amino acid sequence, motif structure, subcellular location, tissue specific distribution and other characteristics of differences and common points, so as to summarize the key residues of the Cd binding target. Then, we explain its special mechanism of action from the molecular structure. In conclusion, this review is expected to provide a reference for the exploration of key amino acid targets of Cd, and lay a foundation for the intelligent design and breeding of crops with high/low Cd accumulation.
Topics: Amino Acids; Cadmium; Inactivation, Metabolic; Metalloproteins; Plants; Stress, Physiological
PubMed: 35163656
DOI: 10.3390/ijms23031734 -
Bioscience Reports Apr 2017Metal ions play pivotal roles in protein structure, function and stability. The functional and structural diversity of proteins in nature expanded with the incorporation... (Review)
Review
Metal ions play pivotal roles in protein structure, function and stability. The functional and structural diversity of proteins in nature expanded with the incorporation of metal ions or clusters in proteins. Approximately one-third of these proteins in the databases contain metal ions. Many biological and chemical processes in nature involve metal ion-binding proteins, aka metalloproteins. Many cellular reactions that underpin life require metalloproteins. Most of the remarkable, complex chemical transformations are catalysed by metalloenzymes. Realization of the importance of metal-binding sites in a variety of cellular events led to the advancement of various computational methods for their prediction and characterization. Furthermore, as structural and functional knowledgebase about metalloproteins is expanding with advances in computational and experimental fields, the focus of the research is now shifting towards design and redesign of metalloproteins to extend nature's own diversity beyond its limits. In this review, we will focus on the computational toolbox for prediction of metal ion-binding sites, metalloprotein design and redesign. We will also give examples of tailor-made artificial metalloproteins designed with the computational toolbox.
Topics: Amino Acid Motifs; Binding Sites; Computational Biology; Databases, Protein; Humans; Metalloproteins; Metals; Models, Molecular; Protein Engineering
PubMed: 28167677
DOI: 10.1042/BSR20160179 -
Biomolecules May 2014The LmbE-like superfamily is comprised of a series of enzymes that use a single catalytic metal ion to catalyze the hydrolysis of various substrates. These substrates... (Review)
Review
The LmbE-like superfamily is comprised of a series of enzymes that use a single catalytic metal ion to catalyze the hydrolysis of various substrates. These substrates are often key metabolites for eukaryotes and prokaryotes, which makes the LmbE-like enzymes important targets for drug development. Herein we review the structure and function of the LmbE-like proteins identified to date. While this is the newest superfamily of metallohydrolases, a growing number of functionally interesting proteins from this superfamily have been characterized. Available crystal structures of LmbE-like proteins reveal a Rossmann fold similar to lactate dehydrogenase, which represented a novel fold for (zinc) metallohydrolases at the time the initial structure was solved. The structural diversity of the N-acetylglucosamine containing substrates affords functional diversity for the LmbE-like enzyme superfamily. The majority of enzymes identified to date are metal-dependent deacetylases that catalyze the hydrolysis of a N-acetylglucosamine moiety on substrate using a combination of amino acid side chains and a single bound metal ion, predominantly zinc. The catalytic zinc is coordinated to proteins via His2-Asp-solvent binding site. Additionally, studies indicate that protein dynamics play important roles in regulating access to the active site and facilitating catalysis for at least two members of this protein superfamily.
Topics: Enzymes; Metalloproteins
PubMed: 24970229
DOI: 10.3390/biom4020527