The handbook is an essential resource for any laboratory studying protein interactions.
Protein Interactions Handbook. The DNA- or RNA-binding function of a protein is localized in discrete conserved domains within its tertiary structure. An individual protein can have multiple repeats of the same nucleic acid binding domain or can have several different domains found within its structure. The identity of the individual domains and their relative arrangement are functionally important within the protein.
Several common DNA binding domains include zinc fingers, helix-turn-helix, helix-loop-helix, winged helix and leucine zipper. Multiple nucleic acid binding domains with a single protein can increase specificity and affinity of the protein for certain target nucleic acid sequences, mediate a change in the topology of the target nucleic acid, properly position other nucleic acid sequences for recognition or regulate the activity of enzymatic domains within the binding protein.
Proteins can bind directly to the nucleic acid or indirectly through other bound proteins, effectively creating a hierarchy of interactions. The strength of these interactions influence which assays or approaches are best for studying complex assembly. Some of these interactions are transient and require stabilization through chemical crosslinking prior to isolation of the complexes.
Specificity is derived from higher order interactions involving nucleoprotein complexes. This interaction is facilitated through the DNA-binding domain s of the transcription factor. In vitro formation of the covalent complex occurs in a two component reaction, i.
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In the cell, before binding the protein recognizes all aspects of the nucleic acid structure, i. In the initial stages of the reaction, the protein interacts with the nucleic acid in a cooperative mode and forms a primary non-covalent, reversible complex. Formation of a tri-substituted phosphate intermediate is possible, which could induce weakening of the adjacent internucleotide bond.
Decay of this intermediate results in breakage of the internucleotide bonds on either side of the phosphorous atom. It also depends on the geometry of the phosphoester bond formed by the hydroxyamino acid residue this was conclusively demonstrated by X-ray crystallography of the DNA-human Topo I and DNA-Cre recombinase intermediate complexes; 23 , Class I proteins are enzymes which combine two activities: cleavage of an internucleotide bond and restoring its formation.
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They belong to the polynucleotide transferase class of enzymes. The covalent complexes are active for polynucleotide transfer to an acceptor molecule. However, our computer analysis A. Barskii and Yu. No energy-rich compounds are necessary for either forming the nucleic acid-protein complex nor restoring the internucleotide bond nor polynucleotide transfer to an acceptor because the interpolymeric complex itself has enough free energy to bring about such reactions.
It appears that cleavage of the internucleotide bond and formation of the covalent complex between nucleic acid and protein take place intramolecularly without the participation of water 23 , 24 , Class I proteins form so-called relaxed complexes with nucleic acids. Denaturation of the protein in the primary complex results in nicking of the nucleotide bond and covalent binding of the protein to the nascent terminal phosphate group of the nucleic acid.
Members of this class of proteins form covalent complexes in both the cytoplasm and the nucleus of the cell. Members of the second class of proteins do not catalyse covalent complex formation and its cleavage.
Special enzymes do. Proteins of this class belong to the so-called terminal proteins and participate in initiation reactions during nucleic acid replication. Energy-rich compounds supply the energy for formation of covalent complexes between Class II proteins and nucleic acids. The compartments associated with covalent complex formation for Class II proteins are cellular membranes.
Initiation of replication of nucleic acids in crude cell membrane complexes has been demonstrated repeatedly 68— Many bacterial plasmids colicinogenic and sex plasmids and drug resistance factors exist as relaxed complexes It should be noted that relaxed complex formation in bacteria depends on the physiological status of the cell 74 and is regulated by catabolite repression It is entirely possible that many eukaryotic circular episomal DNAs can form relaxed complexes similar to those of bacterial complexes.
This has been shown for the plant Ti plasmid In addition, one may readily predict that many extrachromosomal genetic elements [movable genetic elements 77 and cytoplasmic DNAs 78 ] are potential candidates for covalent binding with proteins. According to the mechanisms of formation-disjoining of the covalent complexes, relaxation proteins belong to the class I proteins discussed above.
Natural tight, presumably covalent, complexes of eukaryotic chromosomal DNA and cellular proteins have been under intensive study since the s 8 , 79 , There are two questions crucial to these studies: i what are the functions of the proteins involved? Although the nucleotide-peptides hydrolysed by phosphodiesterases have been isolated from animal 8 , 81 and plant cells 82 , identification of the proteins involved in the linkage remains obscure.
Arrangement of the proteins bound to DNA has mostly been discussed in the context of the structural organization of DNA threading in chromatin 82 , The precise role of covalently bound proteins in chromosomal organization is not completely understood. Another issue is what functional component of chromosomal DNA is a candidate for protein linkage? There are findings that repeated sequence elements Alu family are tightly linked with proteins Summing up the data on covalent complexes of chromosomal DNA with proteins, one can say that proteins with imperfectly identified molecular characteristics and functions are bound via phosphodiester bonds with some DNA sequences 8 , 80 , It seems that bulk isolation of the complexes formed by different proteins makes the structural and functional analyses too complicated.
To date, a few cellular phosphodiester presumably complexes of RNA and protein have been characterized 84— The finding of a p53—5. The function of this complex is presently unknown, but the multifunctional properties of this tumour suppressor protein are well known 88 and references therein. The existence of a new family of nucleoproteins in which nucleic acids are covalently bound to proteins is beyond question.
Proteins & Nucleic Acids
It should be emphasized that the structure of the complex goes hand in hand with its function and that successful studies result from examination of the structure of individual complexes where the partners were identified. To date, one can predict with certainty that the number of unknown nucleic acid-protein covalent complexes is far greater than the number of those known.
The aim of this review has been to provide an overview of the biochemical approaches to be used to obtain convincing evidence regarding the covalent nature of nucleoprotein complexes being investigated now and in the future. I am cordially grateful to Professor A. Bogdanov who, back in the s, involved me in the study of natural covalent complexes of nucleic acids and proteins.
I am indebted to Drs B. Juodka, Z.
Biomaterials for Delivery and Targeting of Proteins and Nucleic Acids - CRC Press Book
Shabarova, Z. Avramova, R. Zanev and D. Werner for helpful discussions. I deeply appreciate Dr B. Semler for critical reading of the manuscript and English language assistance. I am thankful to Dr R. Gumport for critical comments. This work was supported in part by the Russian Foundation for Basic Research grants and and the G. Oxford University Press is a department of the University of Oxford. It furthers the University's objective of excellence in research, scholarship, and education by publishing worldwide.
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Close mobile search navigation Article Navigation. Volume Article Contents. In truth, the success of the Human Genome Project was just a first step. Untangling how, when, and why genes are expressed, and how the resulting proteins function alone and together in the human body is key to realizing the vast potential of the user manual that is our DNA, in concert with the molecules that transcribe it and the proteins it encodes.
And since structure relates to function, such research can shed light on various diseases as well as normal processes. NMR spectroscopy can resolve large molecules as well; using a variety of enhancements, such as selective nucleotide labelling, researchers used two-dimensional solid-state NMR spectroscopy to determine the structure of RNA from the extremophile Pyrococcus furiosus.
But NMR spectroscopy can be used to do much more than determine the structure of a protein or nucleic acid. Electron paramagnetic resonance EPR spectroscopy has also found uses in protein and nucleic acid research.