12 Pancreatic Ribonuclease

Blackburn, Peter; Moore, Stanford

doi:10.1016/s1874-6047(08)60284-x

Cited by 225 publications

(146 citation statements)

References 536 publications

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“…The ribonucleases (RNAses) are a class of enzymes that very effectively catalyze the cleavage of phosphodiester bonds in RNA (Blackburn & Moore, 1982 ;Perreault & Anslyn, 1997 ;Raines, 1998 ;Richards & Wyckoff, 1971) (for instance, RNAse A, in particular, provides an estimated 15 orders of magnitude rate acceleration). RNAse A has been one of the most studied of all enzymes, reflecting in part the fact that this enzyme was among the first enzymes whose structure was solved (see e.g.…”

Section: Ribozymes and Ribonucleasementioning

confidence: 99%

Why nature really chose phosphate

Kamerlin

Sharma

Prasad

et al. 2013

Quart. Rev. Biophys.

332

448

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Phosphoryl transfer plays key roles in signaling, energy transduction, protein synthesis, and maintaining the integrity of the genetic material. On the surface, it would appear to be a simple nucleophile displacement reaction. However, this simplicity is deceptive, as, even in aqueous solution, the low-lying d-orbitals on the phosphorus atom allow for eight distinct mechanistic possibilities, before even introducing the complexities of the enzyme catalyzed reactions. To further complicate matters, while powerful, traditional experimental techniques such as the use of linear free-energy relationships (LFER) or measuring isotope effects cannot make unique distinctions between different potential mechanisms. A quarter of a century has passed since Westheimer wrote his seminal review, ‘Why Nature Chose Phosphate’ (Science 235 (1987), 1173), and a lot has changed in the field since then. The present review revisits this biologically crucial issue, exploring both relevant enzymatic systems as well as the corresponding chemistry in aqueous solution, and demonstrating that the only way key questions in this field are likely to be resolved is through careful theoretical studies (which of course should be able to reproduce all relevant experimental data). Finally, we demonstrate that the reason that naturereallychose phosphate is due to interplay between two counteracting effects: on the one hand, phosphates are negatively charged and the resulting charge-charge repulsion with the attacking nucleophile contributes to the very high barrier for hydrolysis, making phosphate esters among the most inert compounds known. However, biology is not only about reducing the barrier to unfavorable chemical reactions. That is, the same charge-charge repulsion that makes phosphate ester hydrolysis so unfavorable also makes it possible to regulate, by exploiting the electrostatics. This means that phosphate ester hydrolysis can not only be turned on, but also be turned off, by fine tuning the electrostatic environment and the present review demonstrates numerous examples where this is the case. Without this capacity for regulation, it would be impossible to have for instance a signaling or metabolic cascade, where the action of each participant is determined by the fine-tuned activity of the previous piece in the production line. This makes phosphate esters the ideal compounds to facilitate life as we know it.

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Section: Ribozymes and Ribonucleasementioning

confidence: 99%

Why nature really chose phosphate

Kamerlin

Sharma

Prasad

et al. 2013

Quart. Rev. Biophys.

332

448

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“…Of the 123 amino acids in human angiogenin 43 (35%) are identical with those at the corresponding positions in human pancreatic RNase (Beintema et al, 1984) (Table II) and another 41 either are identical with residues in other pancreatic RNases (Blackburn & Moore, 1982;Beintema et al, 1985) or are conservative replacements, constituting an overall homology of 68%.…”

Section: Ribonuclease Homologymentioning

confidence: 99%

Human angiogenin, an organogenic protein

Riordan

Vallee²

1988

Br J Cancer

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Angiogenin is a 14 kD protein, initially isolated as a tumour-cell secreted product but subsequently found to be a normal constituent of human plasma. It is a potent inducer of blood vessel formation on the chorioallantoic membrane of the chick embryo. Chemical characterization of the protein reveals a remarkable homology to the pancreatic ribonuclease family and has led to the identification of a unique ribonucleolytic activity for angiogenin. It is a particularly potent inhibitor of in vitro protein synthesis. Treatment with placental ribonuclease inhibitor abolishes the biological and enzymatic activities of angiogenin, an effect with important mechanistic, physiological and pharmacologic implications.

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“…Paradigmatic examples are bovine seminal RNase (BSRNase) 1,2 and RNase from oocytes of Rana pipiens [3][4][5] (commercial trademark of Alfacell, USA, onconase (ONC)), both belonging to the pancreaticlike RNase family, whose prototype is the bovine pancreatic enzyme (RNase A). 6,7 This unusual biological activity is critically dependent on the ribonucleolytic activity 8 and on the ability of these molecules to reach the cellular cytosol and to degrade RNA by evading the action of the ribonuclease inhibitor (RI). 9,10 It is currently accepted that RI, which binds most RNase A family members with femtomolar affinities, plays an important role in the protection of host cells from endogenous RNases.…”

Section: Introductionmentioning

confidence: 99%

Structural features for the mechanism of antitumor action of a dimeric human pancreatic ribonuclease variant

et al. 2008

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A specialized class of RNases shows a high cytotoxicity toward tumor cell lines, which is critically dependent on their ability to reach the cytosol and to evade the action of the ribonuclease inhibitor (RI). The cytotoxicity and antitumor activity of bovine seminal ribonuclease (BSRNase), which exists in the native state as an equilibrium mixture of a swapped and an unswapped dimer, are peculiar properties of the swapped form. A dimeric variant (HHP2-RNase) of human pancreatic RNase, in which the enzyme has been engineered to reproduce the sequence of BSRNase helix-II (Gln28fiLeu, Arg31fiCys, Arg32fiCys, and Asn34fiLys) and to eliminate a negative charge on the surface (Glu111fiGly), is also extremely cytotoxic. Surprisingly, this activity is associated also to the unswapped form of the protein. The crystal structure reveals that on this molecule the hinge regions, which are highly disordered in the unswapped form of BSRNase, adopt a very well-defined conformation in both subunits. The results suggest that the two hinge peptides and the two Leu28 side chains may provide an anchorage to a transient noncovalent dimer, which maintains Cys31 and Cys32 of the two subunits in proximity, thus stabilizing a quaternary structure, similar to that found for the noncovalent swapped dimer of BSRNase, that allows the molecule to escape RI and/or to enhance the formation of the interchain disulfides.Keywords: crystal structure; antitumor agents; ribonuclease; 3D-domain swapping; dimers Abbreviations: BSRNase, bovine seminal ribonuclease; des(1-7)HP-RNase, human pancreatic ribonuclease in which the first seven residues have been deleted; HHP-RNase, covalent dimeric variant of human pancreatic ribonuclease in which four residues at the helix-II region are mutated according to BSRNase sequence (Gln28!Leu, Arg31!Cys, Arg32!Cys, and Asn34!Lys); HHP2-RNase, HHP-RNase with an additional mutation (Glu111!Gly); HP-RI, structure of human pancreatic ribonuclease complexed with ribonuclease inhibitor; HP-RNase, human pancreatic ribonuclease; MxM-BSRNase, dimeric form of BSRNase in which the chains swap their N-terminal tails; M¼M-BSRNase, unswapped dimer of BSRNase; M¼M-HHP2, unswapped dimer of HHP2-RNase; NCD-BSRNase, noncovalent swapped form of BSRNase; ONC, Onconase; PDB, Protein Data Bank; PM7, human pancreatic ribonuclease in which residues 1-21 are replaced by the corresponding residues of BSRNase; PM8, N-terminal swapped dimer of a variant of human pancreatic ribonuclease; RI, ribonuclease inhibitor; RMSD, root-mean-square deviation.Grant sponsor: Ministero dell'Istruzione, dell'Università e della Ricerca; Grant number: FIRB RBNE03B8KK and PRIN2007.

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12 Pancreatic Ribonuclease

Cited by 225 publications

References 536 publications

Why nature really chose phosphate

Why nature really chose phosphate

Human angiogenin, an organogenic protein

Structural features for the mechanism of antitumor action of a dimeric human pancreatic ribonuclease variant

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