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Nearest-neighbor parameters for predicting DNA duplex stability in diverse molecular crowding conditions.

ABSTRACT: The intracellular environment is crowded and heterogeneous. Although the thermodynamic stability of nucleic acid duplexes is predictable in dilute solutions, methods of predicting such stability under specific intracellular conditions are not yet available. We recently showed that the nearest-neighbor model for self-complementary DNA is valid under molecular crowding condition of 40% polyethylene glycol with an average molecular weight of 200 (PEG 200) in 100 mM NaCl. Here, we determined nearest-neighbor parameters for DNA duplex formation under the same crowding condition to predict the thermodynamics of DNA duplexes in the intracellular environment. Preferential hydration of the nucleotides was found to be the key factor for nearest-neighbor parameters in the crowding condition. The determined parameters were shown to predict the thermodynamic parameters (?H°, ?S°, and ?G°₃₇) and melting temperatures (T _m) of the DNA duplexes in the crowding condition with significant accuracy. Moreover, we proposed a general method for predicting the stability of short DNA duplexes in different cosolutes based on the relationship between duplex stability and the water activity of the cosolute solution. The method described herein would be valuable for investigating biological processes that occur under specific intracellular crowded conditions and for the application of DNA-based biotechnologies in crowded environments.

SUBMITTER: Ghosh S

PROVIDER: S-EPMC7321957 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Nearest-neighbor parameters for predicting DNA duplex stability in diverse molecular crowding conditions.

Ghosh Saptarshi S Takahashi Shuntaro S Ohyama Tatsuya T Endoh Tamaki T Tateishi-Karimata Hisae H Sugimoto Naoki N

Proceedings of the National Academy of Sciences of the United States of America 20200610 25

The intracellular environment is crowded and heterogeneous. Although the thermodynamic stability of nucleic acid duplexes is predictable in dilute solutions, methods of predicting such stability under specific intracellular conditions are not yet available. We recently showed that the nearest-neighbor model for self-complementary DNA is valid under molecular crowding condition of 40% polyethylene glycol with an average molecular weight of 200 (PEG 200) in 100 mM NaCl. Here, we determined nearest ...[more]

PMID: 32522884

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Project description:Quantifying interactions in DNA microarrays is of central importance for a better understanding of their functioning. Hybridization thermodynamics for nucleic acid strands in aqueous solution can be described by the so-called nearest-neighbor model, which estimates the hybridization free energy of a given sequence as a sum of dinucleotide terms. Compared with its solution counterparts, hybridization in DNA microarrays may be hindered due to the presence of a solid surface and of a high density of DNA strands. We present here a study aimed at the determination of hybridization free energies in DNA microarrays. Experiments are performed on custom Agilent slides. The solution contains a single oligonucleotide. The microarray contains spots with a perfect matching complementary sequence and other spots with one or two mismatches: in total 1006 different probe spots, each replicated 15 times per microarray. The free energy parameters are directly fitted from microarray data. The experiments demonstrate a clear correlation between hybridization free energies in the microarray and in solution. The experiments are fully consistent with the Langmuir model at low intensities, but show a clear deviation at intermediate (non-saturating) intensities. These results provide new interesting insights for the quantification of molecular interactions in DNA microarrays. Keywords: experiment to assess technical aspects of hybridization in microarrays The study consists of four experiments. Each experiments consists of one slide with eight identical microarrays which were hybridized with an identical solution but at a different concentration. Per experiment a single target sequence in solution was used for hybridization: target1 to target4 (Table 1 main manuscript or description of samples below). Each target oligo was bought in duplicate, referred to as oligos a and b. In experiment 1 and 2 a fragmentation step (f) was applied. More details can be found in the main manuscript especially Table 3.

Dataset Information

Nearest-neighbor parameters for predicting DNA duplex stability in diverse molecular crowding conditions.

Publications

Nearest-neighbor parameters for predicting DNA duplex stability in diverse molecular crowding conditions.

Similar Datasets

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