Nucleic acids protein sequence translation

Whereas DNA is mostly located in the nucleus of cells in higher organisms (with some also in mitochondria and in plant chloroplasts), RNA comes in three major and distinct forms, each of which plays a crucial role in protein biosynthesis in the cytoplasm. These are, respectively, ribosomal RNA (rRNA), which represents two-thirds of the mass of the ribosome, messenger RNA (mRNA), which encodes the information for the sequence of proteins, and transfer RNAs (tRNAs) which serve as adaptor molecules, allowing the 4-letter code of nucleic acids to be translated into the 20-letter code of proteins. These latter molecules contain a substantial number of modified bases, which are introduced enzymatically. 

Today, the evolution of genes, programmed cell death (apoptosis), and the action of messenger RNA (mRNA) are three major targets of research. mRNA contains the blueprint for every protein in the body. It is transcribed from a DNA template, and carries information to ribosomes, the sites of protein synthesis. The sequences of nucleic acid polymers are translated by transfer RNA (tRNA) into amino acid polymers. tRNA recognizes the three-nucleotide sequences that encode each amino acid. Ribosomal RNA directs the ribosome s production of proteins. Codons carry the messages that terminate protein synthesis. 

If an antibody to the protein of interest is available, it is sometimes possible to use vector sequences, eg, the beta-galactosidase promoter sequence, to direct the transcription of the passenger DNA into messenger RNA and the translation of that mRNA into protein which can be recognized by the antibody. Although this method is somewhat less reHable than the use of nucleic acid probes, specialized vectors are available for this purpose. 

Molecular biology involves the study of the major macromolecules, DNA, RNA, and protein. The central dogma ofmolecular biology is illustrated in Fig. 2. The central dogma shows the relationship among the macromolecules in the processes of transcription and translation. Figure 2 also gives the relationship between immunoelectron microscopy and in situ hybridization. In situ hybridization allows one to localize a specific nucleic acid sequence. Immunoelectron microscopy is an essential component to the technique of in situ hybridization when applied at the EM level. 

A selective method of preventing the expression of adhesion molecules or cytokines is the use of antisense oligonucleotides. These oligonucleotides are short sequences of nucleic acids complementary to mRNA sequences of specific proteins of interest. If delivered to the cytoplasmic compartment of cells these oligonucleotides are able to form a complex with their target mRNA. In this way the translation of mRNA into protein by ribosomes is inhibited. The subsequent mRNA degradation by RNAse H results in reduced expression of the protein (see also Chapter 5 for a description of antisense ohgonucleotides as therapeutic modalities). 

Most of the genetic information stored in the genome codes for the amino acid sequences of proteins. For these proteins to be expressed, a text in nucleic acid language therefore has to be translated into protein language. This is the origin of the use of the term translation to describe protein biosynthesis. The dictionary used for the translation is the genetic code. 

The translating of a sequence of DNA nucleotides into a sequence of amino acids—in other words, into the manufacture of a protein—involves many intricate cellular mechanisms that are still the subject of much research. The overall process, however, is conceptually straightforward, involving two steps— transcription followed by translation. These steps are mediated by the other major nucleic acid, RNA, of which there are three forms—messenger RNA (rnRNA), ribosomal RNA (rRNA), and transfer RNA (tRNA). 

When predicting consensus structures, be aware that the quality of the prediction is limited by the quality of the input alignment. Nucleic acid alignments are generally more error-prone than protein alignments. For coding sequences it is sometimes preferable to back-translate a protein alignment. 

Transfer RNA (tRNA) molecules mediate translation of the nucleic acid genetic code into the amino acid building blocks of proteins, thus ensuring the survivability of cells. The dynamic properties of tRNA molecules are crucial to their functions in both activity and specificity. This chapter summarizes two methods that have been recently developed or improved upon previous protocols to introduce fluorophores to site-specific positions in tRNA. One method enables incorporation of fluorophores carrying a primary amine (such as proflavin or rhodamine) to dihydrouridine (D) residues in the tRNA tertiary core, and a second method enables incorporation of pyrroloC and 2-aminopurine to positions 75 and 76, respectively, of the CCA sequence at the 3 end. These site-specific fluorophore labeling methods utilize tRNA transcripts as the... 

Human apoA-I is a major constituent of HDL, with an Mr of approximately 28,300, calculated from the known primary structure (Bl, B43). ApoA-I is initially synthesized as a 267-amino-acid precursor protein, pre-pro-apoA-I (G25, G26), containing an 18-amino-acid prepeptide and a 6-amino-acid propeptide [determined by nucleic acid sequence analysis of cloned apoA-I (L6), and by isolating the primary translation product of human intestinal apoA-I mRNA (G25)]. 

Finally, novel nucleic acid catalysts have also been selected from random sequence pools (reviewed in Ref. 19). Joyce and co-workers have manipulated the function of the Group I self-splicing ribozyme, selecting variants that can utilize calcium or cleave DNA from partially randomized pools [20,21], Lorsch and Szostak [22] selected a polynucleotide kinase ribozyme from a completely random sequence pool that flanked a previously selected ATP binding site. Many of the novel ribozymes can catalyze reactions that are relevant to protein biosynthesis, bolstering arguments that translation may have arisen in a putative RNA world. For example, Lohse and Szostak [23] have selected ribozymes that can carry out an acyl transfer reaction, while Illangasekare et al. [24] have isolated a... 

---