Three-dimensional domain-swapped oligomers of ribonuclease A: identification of a fifth tetramer, pentamers and hexamers, and detection of trace heptameric, octameric and nonameric species

Gotte, Giovanni; Laurents, Douglas V.; Libonati, Massimo

doi:10.1016/j.bbapap.2005.10.011

Cited by 44 publications

(99 citation statements)

References 19 publications

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“…Each value shown is the mean of three measures from three independent experiments ( s.d. enzyme active site, the higher the ability of the RNase to degrade dsRNA (17,22,24,27,32,45). It has to be recalled here that the preservation of enzymatic activity also seems to be mandatory for a ribonuclease to be cytotoxic or, generally, biologically active (24,30,33,56).…”

Section: Table 3 Enzymatic Activities Of the Various Rnase A Monomermentioning

confidence: 94%

“Zero-Length” Dimers of Ribonuclease A: Further Characterization and No Evidence of Cytotoxicity

et al. 2010

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"Zero-length" dimers of ribonuclease A, a novel type of dimers formed by two RNase A molecules bound to each other through a zero-length amide bond [Simons, B. L., et al. (2007) Proteins 66, 183-195], were further characterized and tested for their possible in vitro cytotoxic activity. Results obtained are the following. Besides dimers, also trimers and higher oligomers could be identified among the products of the covalently linking reaction, and the "zero-length" dimers prepared by us appear not to be a unique species. The product was indeed heterogeneous, and results obtained with two RNase A mutants, E9A and K66A, indicated that amino and carboxyl groups others than those belonging to Lys66 and Glu9 are involved in the amide bond. As for their functional properties, the "zero-length" dimers degrade poly(A).poly(U) (dsRNA) with an activity that increases with the increase of the oligomer's basicity and yeast RNA (ssRNA) with an activity that instead decreases with the increase of oligomer's basicity, which is in agreement with previous data. No cytotoxicity of the RNase A "zero-length" dimers could be evidenced in assays performed with various tumor cells lines; the dimers, instead, become cytotoxic if cationized by conjugation with polyethylenimine (PEI) [Futami et al. (2005) J. Biosci. Bioengin. 99, 95-103]. However, PEI derivatives of RNase A "zero-length" dimers and PEI derivatives of native RNase A resulted to be equally cytotoxic. In other words, protein "dimericity" does not play any role in this case. Moreover, the acquired cytotoxicity does not seem to be specific for tumor cells: PEI-cationized native RNase A was also cytotoxic toward human monocytes.

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Section: Table 3 Enzymatic Activities Of the Various Rnase A Monomermentioning

confidence: 94%

“Zero-Length” Dimers of Ribonuclease A: Further Characterization and No Evidence of Cytotoxicity

et al. 2010

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“…The native monomeric enzyme only degrades single-stranded (ss) RNA and is not cytotoxic, while its artificial dimers and oligomers, either forming through non-covalent DS or covalent bonds, also become active against double-stranded (ds) RNA. 32,92,105,177 Moreover, they can become selectively cytotoxic towards cancer cells both in vitro and in vivo, [178][179][180][181] although some results indicate that cytotoxicity can be dependent also on the type of cell line studied. 35 This acquired cytotoxic power can be considered benign, given the selectivity towards malignant cells, and can be mainly ascribed to the possibility of oligomeric RNases to evade the ribonuclease inhibitor (RI), which is designed to tightly trap monomeric RNases.…”

Section: Bioactivity and Stability Of Protein Oligomersmentioning

confidence: 96%

“…20, panels vii-xi) and possibly more tetramers as well as several other oligomers of higher DP. 30,32,[91][92][93][94] Thus, the folds of RNase A are highly versatile, despite its overall known stability. Its dimers are not exclusively artificial, given that traces of N D are present in a native mixture, 95,96 and that C D has been detected to form and subsequently degrade during protein expression in cells.…”

Section: Self-and Cross-association Through Domain Swapping (Ds)mentioning

confidence: 99%

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A review on protein oligomerization process

Liu

2015

Int. J. Precis. Eng. Manuf.

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Enzymes or proteins are commonly present in oligomeric forms for active biological functions. The formation of protein oligomers, either in nature or by artificial means, can take place via covalent bonding or through weak-bond-network associations. Disulfide bonds are more common in forming covalent protein oligomers. Covalent bound protein oligomers usually have additional elements introduced, while proteins associated through weak-bond-networks usually do not involve any additional bound chemical groups. Proteins are commonly present in stable forms or folds. Oligomerization can occur when stable proteins unfold, creating exposed interfaces for association interactions. One well-known process of weak-bond-network oligomerization is domain swapping (DS). Separating the two touching/interacting domains creates opportunities for similar interactions with different protein molecules, leading to oligomerization. Both disulfide bonding and DS oligomerization can be dynamic (or reversible) at least at low Degree of Polymerizations (DPs). The dynamic nature of the protein oligomerization is important for bioactivity control. The morpheein model and the polymorph model are thus important in quantifying enzyme catalyzed biotransformations. Disulfide bonding and DS can act together to form supramolecules. The formation of supramolecules, such as amyloids, in nature can be benign or harmful. Industrial applications of protein oligomerization exploit the special functions /properties of the resultant supramolecules, such as multiple biofunctions, niche structure, etc..

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“…The structure of the minor trimer has been determined and shows 3D domain swapping only via the C-terminal region, creating a cyclic structure (34). In addition, tetramers or even higher oligomers have been reported for RNase A (35)(36)(37)(38), and four different structural models for the tetramer have been proposed: two linear ones, a propeller-like structure, and a cyclic tetramer (29,35). Unfortunately, as for the major trimer, no atomic structures for any of these higher oligomers have been reported.…”

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confidence: 99%

Different 3D domain-swapped oligomeric cyanovirin-N structures suggest trapped folding intermediates

Koharudin

Liu

Gronenborn

2013

Proc. Natl. Acad. Sci. U.S.A.

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Although it has long been established that the amino acid sequence encodes the fold of a protein, how individual proteins arrive at their final conformation is still difficult to predict, especially for oligomeric structures. Here, we present a comprehensive characterization of oligomeric species of cyanovirin-N that all are formed by a polypeptide chain with the identical amino acid sequence. Structures of the oligomers were determined by X-ray crystallography, and each one exhibits 3D domain swapping. One unique 3D domainswapped structure is observed for the trimer, while for both dimer and tetramer, two different 3D domain-swapped structures were obtained. In addition to the previously identified hinge-loop region of the 3D domain-swapped dimer, which resides between strands β5 and β6 in the middle of the polypeptide sequence, another hinge-loop region is observed between strands β7 and β8 in the structures. Plasticity in these two regions allows for variability in dihedral angles and concomitant differences in chain conformation that results in the differently 3D domain-swapped multimers. Based on all of the different structures, we propose possible folding pathways for this protein. Altogether, our results illuminate the amazing ability of cyanovirin-N to proceed down different folding paths and provide general insights into oligomer formation via 3D domain swapping.CV-N | NMR spectroscopy | protein folding | X-ray structures

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Three-dimensional domain-swapped oligomers of ribonuclease A: identification of a fifth tetramer, pentamers and hexamers, and detection of trace heptameric, octameric and nonameric species

Cited by 44 publications

References 19 publications

“Zero-Length” Dimers of Ribonuclease A: Further Characterization and No Evidence of Cytotoxicity

“Zero-Length” Dimers of Ribonuclease A: Further Characterization and No Evidence of Cytotoxicity

A review on protein oligomerization process

Different 3D domain-swapped oligomeric cyanovirin-N structures suggest trapped folding intermediates

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