New Insight into Secreted Ribonuclease Structure: Binase Is a Natural Dimer

Dudkina, Elena; Kayumov, Airat R.; Ulyanova, Vera; Ilinskaya, О. N.

doi:10.1371/journal.pone.0115818

Cited by 20 publications

(27 citation statements)

References 45 publications

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“…The immunoblot analysis showed that anti-binase antibodies interact with low order oligomers of balifase too ( Figure 4(d) ). Therefore, we can conclude that balifase can exist in oligomeric forms in solution but unlike binase, not under natural conditions of biosynthesis [ 25 ].…”

Section: Resultsmentioning

confidence: 99%

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Extracellular Ribonuclease fromBacillus licheniformis(Balifase), a New Member of the N1/T1 RNase Superfamily

Sokurenko

Nadyrova

Ulyanova

et al. 2016

BioMed Research International

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The N1/T1 RNase superfamily comprises enzymes with well-established antitumor effects, such as ribotoxins secreted by fungi, primarily by Aspergillus and Penicillium species, and bacterial RNase secreted by B. pumilus (binase) and B. amyloliquefaciens (barnase). RNase is regarded as an alternative to classical chemotherapeutic agents due to its selective cytotoxicity towards tumor cells. New RNase with a high degree of structural similarity with binase (73%) and barnase (74%) was isolated and purified from Bacillus licheniformis (balifase, calculated molecular weight 12421.9 Da, pI 8.91). The protein sample with enzymatic activity of 1.5 × 106 units/A280 was obtained. The physicochemical properties of balifase are similar to those of barnase. However, in terms of its gene organization and promoter activity, balifase is closer to binase. The unique feature of balifase gene organization consists in the fact that genes of RNase and its inhibitor are located in one operon. Similarly to biosynthesis of binase, balifase synthesis is induced under phosphate starvation; however, in contrast to binase, balifase does not form dimers under natural conditions. We propose that the highest stability of balifase among analyzed RNase types allows the protein to retain its structure without oligomerization.

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Section: Resultsmentioning

confidence: 99%

“…To estimate in-gel RNase activity of proteins we performed zymography analysis as described in [ 25 ]. Proteins were separated in 15% polyacrylamide gel with 0.1% SDS (SDS-PAGE) [ 23 ].…”

Section: Methodsmentioning

confidence: 99%

Extracellular Ribonuclease fromBacillus licheniformis(Balifase), a New Member of the N1/T1 RNase Superfamily

Sokurenko

Nadyrova

Ulyanova

et al. 2016

BioMed Research International

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“…It is known that peak asymmetry could indicate the presence of different oligomeric forms of enzyme in the sample [33]. Moreover, it was found that binase forms very stable dimers which do not dissociate during SDS/PAGE [25]. These facts denoted the possibility of balnase to dimerize.…”

Section: Discussionmentioning

confidence: 99%

Three‐step procedure for preparation of pure Bacillus altitudinis ribonuclease

et al. 2016

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Ribonucleases are considered as promising tools for anticancer treatment due to their selective cytotoxicity against tumor cells. We investigated a new RN ase from Bacillus altitudinis termed BALNASE ( B . altitudinis RN ase). Balnase is a close homolog of the well‐known cytotoxic binase, differing by only one amino acid residue: nonpolar hydrophobic alanine at position 106 in the balnase molecule is replaced by a polar uncharged threonine in binase. The most exciting question is how the physico‐chemical properties and biological effects of RN ase might be changed by A106T substitution. Here, we have developed a chromatography‐based rapid and modern technique for the purification of this new RN ase which allowed us to get a protein sample of high quality with specific activity of 1.2 × 10 6 units in preparative amounts, suitable for further investigation of its biological properties.

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“… Ribonuclease binase from B.pumilus was cleaved by trypsin and the cleavage fragments were analysed on HPLC LC-MS/MS system (Bruker, Germany) 60 . The numbers denote determined oligopeptide masses.…”

Section: Figurementioning

confidence: 99%

Statistical prediction of protein structural, localization and functional properties by the analysis of its fragment mass distributions after proteolytic cleavage

Bogachev

Kayumov

Markelov

et al. 2016

Sci Rep

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Structural, localization and functional properties of unknown proteins are often being predicted from their primary polypeptide chains using sequence alignment with already characterized proteins and consequent molecular modeling. Here we suggest an approach to predict various structural and structure-associated properties of proteins directly from the mass distributions of their proteolytic cleavage fragments. For amino-acid-specific cleavages, the distributions of fragment masses are determined by the distributions of inter-amino-acid intervals in the protein, that in turn apparently reflect its structural and structure-related features. Large-scale computer simulations revealed that for transmembrane proteins, either α-helical or β -barrel secondary structure could be predicted with about 90% accuracy after thermolysin cleavage. Moreover, 3/4 intrinsically disordered proteins could be correctly distinguished from proteins with fixed three-dimensional structure belonging to all four SCOP structural classes by combining 3–4 different cleavages. Additionally, in some cases the protein cellular localization (cytosolic or membrane-associated) and its host organism (Firmicute or Proteobacteria) could be predicted with around 80% accuracy. In contrast to cytosolic proteins, for membrane-associated proteins exhibiting specific structural conformations, their monotopic or transmembrane localization and functional group (ATP-binding, transporters, sensors and so on) could be also predicted with high accuracy and particular robustness against missing cleavages.

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New Insight into Secreted Ribonuclease Structure: Binase Is a Natural Dimer

Cited by 20 publications

References 45 publications

Extracellular Ribonuclease fromBacillus licheniformis(Balifase), a New Member of the N1/T1 RNase Superfamily

Extracellular Ribonuclease fromBacillus licheniformis(Balifase), a New Member of the N1/T1 RNase Superfamily

Three‐step procedure for preparation of pure Bacillus altitudinis ribonuclease

Statistical prediction of protein structural, localization and functional properties by the analysis of its fragment mass distributions after proteolytic cleavage

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