Contents
CHAPTER 5: CHAPTER 5: STUDIES ON THE AGGREGATION INDUCED FLUORESCENCE EMISSION PROPERTY OF PYRENYLAMIDO
1.1. Introduction
1.1.1. Nucleosides, Nucleotides and Nucleic Acids
Nucleic acids are one of the most essential building blocks of living cells. Chemists and biochemists recognize nucleic acids as the center of importance in biological systems.
Structurally, nucleic acids exist in long polymeric structures consist of nucleotides. Even though their compositions are simple nucleotide monomers, these biopolymers take an active part in various complex cellular functions. For example, deoxyribonucleic acid (DNA) stores, access and replicates genetic information as a linear nucleotide code. On the other hand, ribonucleic acid (RNA) transports genetic information from DNA to the ribosome which ultimately leads to protein synthesis. The role of nucleic acids as a genetic carrier was not realized until about 70 years ago. During last half century, numerous experiments and investigations were performed significantly on the structure of DNA, 1-7 structure-function relationships between DNA and RNA 8, 9 and fundamental processes like DNA replication, RNA transcription and protein synthesis.10-15 The observations and results of these investigations lead to modern molecular biology and form the basis of nucleic acids as a genetic carrier. The transportation of genetic information from DNA into RNA and then to a protein known as the central dogma of molecular biology originally put forward by Francis Crick (Figure 1.1).16, 17
Figure 1.1. The central dogma of molecular biology
According to the theory of central dogma, an individual DNA molecule serves as templates for complementary DNA strands during the replication process, while as complementary RNA molecules during the transcription process. Sequentially, the RNA molecules serve as fingerprints for the arrangements of amino acids by ribosomes during the process of translation.
The principle of this paradigm have withstood the test of time and experimentation and continues to represent as guiding principle for molecular biologists involving in all areas of basic biological, biomedical and genetic research. However, certain additions are necessary which are based on observations and discovery of some unusual transcriptions. These include the occurrence of RNA synthesis and RNA-directed DNA synthesis (reverse transcription), as found in some viruses and plant species.18 Later on, various experiments and their findings indicate the involvement of RNA in more complex biological activities than DNA.19, 20
1.1.2. Structure of Nucleic Acids
Deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) are the two types of nucleic acids present in living cells as mentioned before. Both are polymers of repeating subunits called nucleotides. The arrangement of nucleotide subunits in the primary structure of a nucleic acid responsible for the flow of genetic information within a cell. Each nucleotide monomer consists of (i) a pentose (5 carbon) sugar, (ii) a base, which is essentially a cyclic nitrogen-containing compound, and (iii) a phosphate group. There are two types of sugar present in nucleic acids, deoxyribose which is present only in DNA and ribose which is present in RNA. Therefore, the two nucleic acids are named according to the sugars present in them. The only structural difference between these two sugars is the absence of oxygen at 2'-position of deoxyribose sugar (1.004, 1.005, Figure 1.2). The bases present in nucleic acids resemble either a purine ring system or a pyrimidine ring system and termed accordingly as purines and pyrimidines (Figure 1.2). These bases are also called as nucleobases as they primarily found in nucleic acids. In DNA, four different bases are found naturally: adenine (A), guanine (G), cytosine (C) and thymine (T). The first two are derivatives of purine whereas the other two are derivatives of pyrimidine. In RNA, we find the same first three nucleobases as in DNA but instead of thymine, another pyrimidine derivative uracil (U) is present as a fourth nucleobase.
In addition to these major bases, there is also a large range of minor bases which occur less frequently than others. A wide variety of modified nucleobases are found in RNAs, whereas the DNA of eukaryotes consists of simply modified nucleobases involving the methylation of
the C5-position of cytosine or the exocyclic amino group of adenine. These modifications support a mechanism of regulation and expression of individual genes at the DNA level in eukaryotes. 21 The third component of a nucleotide monomer is a phosphate group which is derived from phosphoric acid (1.006, Figure 1.2).
Figure 1.2. Components of a nucleotide.
Figure 1.3. Natural nucleosides and their ring numbering system.
The structure composed by any one of the nucleobases with either one of the two sugar moieties via a glycosidic bond is known as nucleoside (Figure 1.3). When the sugar moiety is ribose then we have ribonucleoside, on the other hand, if it is deoxyribose then we have a deoxyribonucleoside. Accordingly, nucleosides with individual bases and ribose sugar are termed as adenosine, guanidine, cytidine, thymidine, uridine respectively. On the hand, nucleosides with individual bases and deoxyribose sugar are termed as deoxyadenosine, deoxyguanidine, deoxycytidine, deoxythymidine, and deoxyuridine respectively. Structurally, a nucleotide is a nucleoside phosphate. These nucleotide monomers are connected together to
form a polymeric structure by 3', 5'-phosphodiester bonds leading to the primary structure nucleic acids (Figure 1.4).
Figure 1.4. The molecular structure of Nucleotides, DNA and RNA.
In addition, one or two additional phosphates can be inserted into the first phosphate group of a nucleoside molecule via a pyrophosphate linkage. The nucleosides with one phosphate group are called nucleoside monophosphates (NMPs), those with two and three phosphate groups are called nucleoside diphosphates (NDPs) and nucleoside triphosphates (NTPs) respectively. One of the most important nucleobases involved in these compounds is adenine which forms adenosine mono, di and triphosphate molecules (AMP, ADP, and ATP) (Figure 1.5). These molecules are well-known for their vital roles in many biological processes.
Figure 1.5. Adenosine phosphates.
One of the most important structural features of DNA is the specific pairing of nucleobases.
In a natural DNA, adenine always pairs with thymine and cytosine always pairs with guanine and vice versa. This base pairing is specific in DNA and during polymerase-mediated replication, it forms the basis of the genetic alphabet, ultimately leading to the basis of the genetic code.22, 23 However, the genetic alphabets and hence the information stored in them by no means should be restricted to only two base pairs. It became a logical thought among the scientific community that an expanded genetic alphabet could provide us a platform for the encoding of additional information which might be beneficial not only for various in vitro and in vivo applications but also for a variety of biotechnology applications. Toward the expansion of genetic alphabet, an unnatural third base pair can be formed between two identical unnatural nucleotides (self-pairs) or two different unnatural nucleotides (hetero pairs). The incorporation of such unnatural base pairs into DNA would expand the potential of DNA in terms of information and functions such as site-directed labeling of oligonucleotides and in vitro selections with oligonucleotides functionalized with diverse chemical structures.24-26 Thus, the design and synthesis of unnatural base pairs is an interesting research area leading towards the expansion of genetic alphabet which would find widespread applications in biotechnology including the production of unnatural proteins and organisms.27-29
Figure 1.6: Presentation of hydrogen bonding between the DNA bases.