Immobilized smart RNA on graphene oxide nanosheets to specifically recognize and adsorb trace peptide toxins in drinking water

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Abstract

The contaminations of peptide toxins in drinking water lead directly to sickness and even death in both humans and animals. A smart RNA as aptamer is covalently immobilized on graphene oxide to form a polydispersed and stable RNA–graphene oxide nanosheet. RNA–graphene oxide nanosheets can resist nuclease and natural organic matter, and specifically adsorb trace peptide toxin (microcystin-LR) in drinking water. The adsorption data fit the pseudo-second-order kinetics and the Langmuir isotherm model. The adsorption capacity of RNA–graphene oxide nanosheets decreases at extreme pH, temperature, ionic strength and natural organic matter, but it is suitable to adsorb trance pollutants in contaminated drinking water. Compared with other chemical and biological sorbents, RNA–graphene oxide nanosheets present specific and competitive adsorption, and are easily synthesized and regenerated. Aptamer (RNA) covalently immobilized on graphene oxide nanosheets is a potentially useful tool in recognizing, enriching and separating small molecules and biomacromolecules in the purification of contaminated water and the preparation of samples.

Highlights

► We covalently immobilize smart RNA (aptamer) on graphene oxide (GO) nanosheets. ► RNA–GO can resist nuclease and nature organic matter. ► RNA–GO can specially recognizes and adsorbs trace toxins in drinking water. ► RNA–GO can be used in environmental, chemical and biological science.

Introduction

Cyanobacteria species vigorously produce toxic compounds, frequently referred to as cyanotoxins. These toxins are globally recognized as triggering health risks via sources of drinking water [1]. The high chemical stability and water solubility of cyanotoxins lead to a long persistence of contamination in sources of drinking water. The most prevalent of cyanotoxins is peptide microcystin-LR that has led to serious sicknesses and deaths in people and animals [1]. Recent studies have exhibited trace microcystin-LR (ng L−1 μg L−1) in drinking water, and confirmed that very low concentrations (less than 1 μg L−1) significantly interrupted cellular processes [2]. Therefore, an effective adsorbent is crucial for scientists to detect and remove trace microcystin-LR in drinking water.

Concern about the global occurrence of biological toxin contamination has prompted the development of adsorption and removal of toxins in drinking water. Recent advances in nanomaterials and biomaterials have attractive properties for the adsorption and removal of peptide toxins. Nanomaterials, such as silver and titanium dioxide nanoparticles, have been used to adsorb and remove microcystin-LR from water [3]. However, several challenges exist in the efficient application of nanomaterials, for example, aggregation, nonspecific adsorption, secondary contamination and toxicity [4]. In addition, adsorption onto geosorbents/coating by natural organic matter (NOM) reduces the efficiency of nanomaterials [5]. Biomaterials, such as bacteria and peat, were used to adsorb toxins in water [6], [7], as well as there should be increased attention paid to secondary contamination and the efficiency at low concentrations (ng L−1) on the adsorption using traditional biomaterials [8]. Antibodies or enzymes have been widely used to detect or adsorb peptide toxins by immunoassays and immunoaffinity [9]. Notably, the complex selection processes and tricky stability of antibodies and enzymes are the bottlenecks of the relevant application.

The above problems could be solved by aptamers. Aptamers, a new class of single-stranded DNA (ssDNA)/RNA molecules, are selected from synthetic nucleic acid libraries for molecular recognition [10], [11], [12]. The smart ssDNA/RNA can recognize various targets molecules, including proteins, nucleic acids, peptides, amino acids, cells, bacterial viruses, organics and small molecules [11], [12]. In contrast to antibodies or enzymes, the smart ssDNA/RNA has the advantages of high affinity, specificity and stability with targets, and easy to prepare and modify at low cost [13]. The bare and supported ssDNA have been used to remove arsenide and mercury in water, respectively [14], [15]. Furthermore, ssDNA was immobilized on sepharose particles to remove trace pharmaceuticals from drinking water [16]. It is known that ssDNA/RNA is unstable in the environment and easy to degrade by nuclease. The effects of NOM, nuclease and other physicochemical conditions could pose serious challenges for the application of the smart ssDNA/RNA, which is still obscure.

Graphene oxide (GO) is a newly discovered nanomaterial possessing a large surface area (2630 m2 g−1) with little toxicity, which is attractive to water purification [17], [18], [19], [20]. Aptamer (ssDNA) noncovalently adsorbed on GO can resist DNase, but ssDNA releases from GO when it contacts targets [21]. Therefore, the noncovalent immobilzation of ssDNA/RNA is not a valid method against nuclease. In this work, smart RNA is covalently immobilized on GO nanosheets to enhance the stability of RNA, and then specifically adsorb trace (ng L−1) microcystin-LR in drinking water. Two points are supposed for RNA–GO: (i) increasing the specific adsorption of GO nanosheets due to the smart RNA modification and (ii) enhancing the stability of RNA owing to the covalent immobilization on GO nanosheets. First, RNA–GO nanosheets are synthesized and then characterized using Fourier transform infrared spectra (FTIR) and atomic force microscopy (AFM) image, as well as the stability of RNA–GO against nuclease is explored by agarose gel electrophoresis. Subsequently, the capacity of recognition of RNA–GO is tested by specifically binding to the similar structure of peptide toxins. Furthermore, the adsorption characteristics and effects of physicochemical conditions (pH, temperature, ionic strength and NOM) are expounded. Finally, RNA–GO as a novel sorbent is compared with other chemical and biological materials.

Section snippets

Materials

The aptamer (RNA) specifically binding to microcystin-LR was originally selected from a random RNA pool (6 × 1014) by Gu and Famulok [22]. The sequence is described as the following: 5′-GGGAGAGACAAGCUUGGGUCCCGGGGUAGGGAUGGGAGGUAUGGAGGGGUCCUUGUUUCCCUCUUGCUCUUCCUAGGAGU-3′

The first and last 20 bases are the primers for the RNA selection, as the italic characters. The two ends of a random RNA have the same sequence as the smart RNA. The middle 40 bases of the random RNA sequence are set with the same

Characteristics of GO and RNA–GO nanosheets

FTIR, AFM and agarose gel electrophoresis were used to expound the characteristics of GO and RNA–GO, as shown in Fig. 2. FTIR was to identify the functional groups attached on GO and RNA–GO, as described in Fig. 2a. GO exhibited a strong band at 1, 750 cm−1 assigned to the Cdouble bondO stretching vibrations from carbonyl and carboxyl groups. The band at 1200 cm−1 assigned to the Csingle bondOH stretching vibrations was not obvious due to the modification of NaOH and chloroacetic acid. The spectrum of GO has a

Conclusions

Aptamer (RNA) was covalently immobilized on graphene oxide to form a polydispersed and stable nanosheet. RNA–graphene oxide can resist nuclease and natural organic matter, and specifically adsorb trace peptide toxins in drinking water. The adsorption data fit well with the pseudo-second-order kinetics and the Langmuir isotherm model. Extreme pH, temperature and ionic strength reduce the adsorption, as well as RNA–GO exhibits specific adsorption, high efficacy, and facile synthesis and

Acknowledgment

This work was financially supported by the National Natural Science Foundation of China as a key project (21037002), the Shanghai Tongji Gao Tingyao Environmental Science & Technology Development Foundation (STGEF), the National Sciences and Engineering Research Council of Canada and the China Scholarship Council.

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