Population shift vs induced fit: The case of bovine seminal ribonuclease swapping dimer

Abstract

Bovine seminal ribonuclease (BS-RNase) is a unique member of the pancreatic-like ribonuclease superfamily. This enzyme exists as two conformational isomers with distinctive biological properties. The structure of the major isomer is characterized by the swapping of the N-terminal segment (MxM BS-RNase). In this article, the crystal structures of the ligand-free MxM BS-RNase and its complex with 2'-deoxycitidylyl(3',5')-2'-deoxyadenosine derived from isomorphous crystals have been refined. Interestingly, the comparison between this novel ligand-free form and the previously published sulfate-bound structure reveals significant differences. In particular, the ligand-free MxM BS-RNase is closer to the structure of MxM BS-RNase productive complexes than to the sulfate-bound form. These results reveal that MxM BS-RNase presents a remarkable flexibility, despite the structural constraints of the interchain disulfide bridges and the swapping of the N-terminal helices. These findings have important implications to the ligand binding mechanism of MxM BS-RNase. Indeed, a population shift rather than a substrate-induced conformational transition may occur in the MxM BS-RNase ligand binding process.

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

“…Thus, the interchain disulphide bonds and the two mutations Gln28!Leu and Asn34!Lys reproduce in M¼M-HHP2 the same helix-II/helix-II interface characteristic of the seminal ribonuclease. [18][19][20] This similarity extends to the two Leu28 side chains, which make stabilizing hydrophobic interactions across the molecular twofold axis. The side chains of Lys34 are completely disordered, as in M¼M-BSRNase and MxM-BSRNase.…”

“…The quaternary association of M¼M-HHP2 is very similar to that of the covalent dimeric forms of BSRNase [18][19][20] : after the superimposition of the core of one subunit, a further rotation of only 8.2 and 7.3 is required to fit the core of the second subunit of M¼M-BSRNase and MxM-BSRNase, respectively. Thus, the interchain disulphide bonds and the two mutations Gln28!Leu and Asn34!Lys reproduce in M¼M-HHP2 the same helix-II/helix-II interface characteristic of the seminal ribonuclease.…”

“…The side chains of Lys34 are completely disordered, as in M¼M-BSRNase and MxM-BSRNase. [18][19][20] The two active sites are very well defined and show minor structural differences [ Fig. 1(B)], probably related to the crystallization pH (6.8), which is close to the pK a of the catalytic histidines.…”

“…[18][19][20] In both isoforms, the dimeric interface is formed by the hinge peptides (residues [16][17][18][19][20][21][22] and the helices (residues [23][24][25][26][27][28][29][30][31][32][33][34] that comprise the four cysteines forming the two interchain disulfide bridges (Cys31-Cys32 0 and Cys32-Cys31 0 ) and the side chains of Leu28 and 28 0 that form stabilizing hydrophobic interactions across the molecular twofold axis. 18,20 In the most abundant swapped form, MxM-BSRNase, the dimeric interface also includes the closed interface arising from the swapped elements. 18,20 As a consequence of this latter contribution, MxM-BSRNase gives rise, upon selective reduction of the intersubunit disulfides, to a metastable noncovalent swapped dimer (NCD-BSRNase), whereas M¼M-BSRNase yields a monomeric derivative.…”

“…18,20 In the most abundant swapped form, MxM-BSRNase, the dimeric interface also includes the closed interface arising from the swapped elements. 18,20 As a consequence of this latter contribution, MxM-BSRNase gives rise, upon selective reduction of the intersubunit disulfides, to a metastable noncovalent swapped dimer (NCD-BSRNase), whereas M¼M-BSRNase yields a monomeric derivative. 17 Although the two dimers present an almost undistinguishable external shape, 21 the cytotoxic action is a peculiar property of MxM-BSRNase.…”