How to Win the Battle with RNase

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Abstract

Because ribose residues carry hydroxyl groups in both the 2′ and 3′ positions, RNA is chemically much more reactive than DNA and is easy prey to cleavage by contaminating RNases—enzymes with various specificities that share the property of hydrolyzing diester bonds linking phosphate and ribose residues. Because RNases are released from cells following lysis and are present on the skin, constant vigilance is required to prevent contamination of glassware and bench tops and the creation of aerosols carrying RNase. The problem is compounded because there is no simple method to inactivate RNases. Because of the presence of intrachain disulfide bonds, many RNases are resistant to prolonged boiling and mild denaturants and are able to refold quickly when denatured. Unlike many DNases, RNases do not require divalent cations for activity and thus cannot be easily inactivated by the inclusion of ethylenediaminetetraacetic acid (EDTA) or other metal ion chelators in buffer solutions. The best way to prevent problems with RNase is to avoid contamination in the first place.

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A structural basis of the interactions between leucine-rich repeats and protein ligands.

B Kobe, J. Deisenhofer (1995)

The leucine-rich repeat is a recently characterized structural motif used in molecular recognition processes as diverse as signal transduction, cell adhesion, cell development, DNA repair and RNA processing. We present here the crystal structure at 2.5 A resolution of the complex between ribonuclease A and ribonuclease inhibitor, a protein built entirely of leucine-rich repeats. The unusual non-globular structure of ribonuclease inhibitor, its solvent-exposed parallel beta-sheet and the conformational flexibility of the structure are used in the interaction; they appear to be the principal reasons for the effectiveness of leucine-rich repeats as protein-binding motifs. The structure can serve as a model for the interactions of other proteins containing leucine-rich repeats with their ligands.

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Crystal structure of porcine ribonuclease inhibitor, a protein with leucine-rich repeats.

B Kobe, J. Deisenhofer (2015)

Ribonuclease inhibitor is a cytoplasmic protein that tightly binds and inhibits ribonucleases of the pancreatic ribonuclease superfamily. The primary sequence of this inhibitor contains leucine-rich repeats (LRRs); these motifs are present in many proteins that participate in protein-protein interactions and have different functions and cellular locations. In vivo, ribonuclease inhibitor may have a role in the regulation of RNA turnover in mammalian cells and in angiogenesis. To define the structural features of LRR proteins and to understand better the nature of the tight interaction of ribonuclease inhibitor with ribonucleases, we have determined the crystal structure of the porcine inhibitor. To our knowledge, this is the first three-dimensional structure of a protein containing LRRs and represents a new class of alpha/beta protein fold. Individual repeats constitute beta-alpha structural units that probably also occur in other proteins containing LRRs. The non-globular shape of the structure and the exposed face of the parallel beta-sheet may explain why LRRs are used to achieve strong protein-protein interactions. A possible ribonuclease-binding region incorporates the surface formed by the parallel beta-sheet and the beta alpha loops.