Ribonuclease inhibitor (RI) is a large (~450 residues, ~49 kDa), acidic (pI ~4.7),
leucine-rich repeat
A leucine-rich repeat (LRR) is a protein structural motif that forms an α/β horseshoe fold. It is composed of repeating 20–30 amino acid stretches that are unusually rich in the hydrophobic amino acid leucine. These tandem repeats commo ...
protein that forms extremely tight complexes with certain
ribonuclease
Ribonuclease (commonly abbreviated RNase) is a type of nuclease that catalyzes the degradation of RNA into smaller components. Ribonucleases can be divided into endoribonucleases and exoribonucleases, and comprise several sub-classes within the ...
s. It is a major cellular protein, comprising ~0.1% of all cellular protein by weight, and appears to play an important role in regulating the lifetime of
RNA.
RI has a surprisingly high
cysteine content (~6.5%, cf. 1.7% in typical proteins) and is sensitive to oxidation. RI is also rich in
leucine
Leucine (symbol Leu or L) is an essential amino acid that is used in the biosynthesis of proteins. Leucine is an α-amino acid, meaning it contains an α-amino group (which is in the protonated −NH3+ form under biological conditions), an α- c ...
(21.5%, compared to 9% in typical proteins) and commensurately lower in other hydrophobic residues, esp.
valine,
isoleucine,
methionine,
tyrosine, and
phenylalanine.
Structure
RI is the classic leucine-rich repeat protein, consisting of alternating
α-helices
The alpha helix (α-helix) is a common motif in the secondary structure of proteins and is a right hand-helix conformation in which every backbone N−H group hydrogen bonds to the backbone C=O group of the amino acid located four residues ear ...
and
β-strands along its backbone. These
secondary structure elements wrap around in a curved, right-handed solenoid that resembles a
horseshoe. The parallel β-strands and α-helices form the inner and outer wall of the horseshoe, respectively. The structure appears to be stabilized by buried
asparagine
Asparagine (symbol Asn or N) is an α-amino acid that is used in the biosynthesis of proteins. It contains an α-amino group (which is in the protonated −NH form under biological conditions), an α-carboxylic acid group (which is in the depro ...
s at the base of each turn, as it passes from α-helix to β-strand. The αβ repeats alternate between 28 and 29 residues in length, effectively forming a 57-residue unit that corresponds to its genetic structure (each
exon codes for a 57-residue unit).
Binding to ribonucleases
The
affinity
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of RI for ribonucleases is among the highest for any
protein-protein interaction; the
dissociation constant
In chemistry, biochemistry, and pharmacology, a dissociation constant (K_D) is a specific type of equilibrium constant that measures the propensity of a larger object to separate (dissociate) reversibly into smaller components, as when a complex f ...
of the RI-
RNase A
Pancreatic ribonuclease family (, ''RNase'', ''RNase I'', ''RNase A'', ''pancreatic RNase'', ''ribonuclease I'', ''endoribonuclease I'', ''ribonucleic phosphatase'', ''alkaline ribonuclease'', ''ribonuclease'', ''gene S glycoproteins'', ''Ceratit ...
complex is in the
femtomolar
Molar concentration (also called molarity, amount concentration or substance concentration) is a measure of the concentration of a chemical species, in particular of a solute in a solution, in terms of amount of substance per unit volume of solu ...
(fM) range under physiological conditions while that for the RI-
angiogenin
Angiogenin (ANG) also known as ribonuclease 5 is a small 123 amino acid protein that in humans is encoded by the ''ANG'' gene. Angiogenin is a potent stimulator of new blood vessels through the process of angiogenesis. Ang hydrolyzes cellular ...
complex is less than 1 fM. Despite this high affinity, RI is able to bind a wide variety of RNases A despite their relatively low
sequence identity
In bioinformatics, a sequence alignment is a way of arranging the sequences of DNA, RNA, or protein to identify regions of similarity that may be a consequence of functional, structural, or evolutionary relationships between the sequences. Ali ...
. Both biochemical studies and
crystallographic structures of RI-RNase A complexes suggest that the interaction is governed largely by
electrostatic interactions, but also involves substantial buried
surface area. RI's affinity for ribonucleases is important, since many ribonucleases have
cytotoxic
Cytotoxicity is the quality of being toxic to cells. Examples of toxic agents are an immune cell or some types of venom, e.g. from the puff adder (''Bitis arietans'') or brown recluse spider (''Loxosceles reclusa'').
Cell physiology
Treating cel ...
and
cytostatic
Cytostasis (cyto – cell; stasis – stoppage) is the inhibition of cell growth and multiplication. Cytostatic refers to a cellular component or medicine that inhibits cell division.
Cytostasis is an important prerequisite for structured multic ...
effects that correlate well with ability to bind RI.
Mammalian RIs are unable to bind certain pancreatic ribonuclease family members from other species. In particular,
amphibian
Amphibians are four-limbed and ectothermic vertebrates of the class Amphibia. All living amphibians belong to the group Lissamphibia. They inhabit a wide variety of habitats, with most species living within terrestrial, fossorial, arborea ...
RNases, such
ranpirnase
Ranpirnase is a ribonuclease enzyme found in the oocytes of the Northern Leopard Frog (''Rana pipiens''). Ranpirnase is a member of the pancreatic ribonuclease (RNase A) protein superfamily and degrades RNA substrates with a sequence preference f ...
and
amphinase
Amphinase is a ribonuclease enzyme found in the oocytes of the Northern leopard frog ''(Rana pipiens)''. Amphinase is a member of the pancreatic ribonuclease protein superfamily and degrades long RNA substrates. Along with ranpirnase, another le ...
from the
Northern leopard frog, escape mammalian RI and have been noted to have differential cytotoxicity against
cancer cells.
See also
*
Guanidinium thiocyanate - a chemical RNase inhibitor.
References
Further reading
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{{Enzyme inhibition
Protein domains
Hydrolase inhibitors
LRR proteins