class-12-molecular-basis-of-inheritance ii neet-aiims
Sites of Protein Synthesis
l Protein synthesis occurs over ribosomes and, therefore, also called protein factories. They are minute spherical nucleoprotein entities which are devoid of covering membranes. The major components of ribosomes are RNA and proteins. In the entire ribosome, RNA makes 66% part in prokaryotes and 60% in eukaryotes. There is no lipid content in the ribosomes. Ribosomes may form a helical group of 5-20, called polyribosomes or polysomes (Rich, 1963). The adjacent ribosome of a polyribosome is about 340 A° apart. Ribosomes are joined to the mRNA molecule. Each ribosome has two subunits, small and large.
The smaller subunit is the 40S and the larger one is 60S in eukaryotes while in prokaryotes these are 30S and 50S, respectively. The larger subunit (the 50S) in prokaryotes consists of a 23S rRNA molecule, a 5S rRNA molecule, and 31 ribosomal proteins. In the eukaryotic equivalents (the 60S), a 28 S rRNA molecule is accompanied by a 5.8 S rRNA and 5 S rRNA molecule and 49 proteins. The smaller prokaryotic subunit (the 30S) consists of a 16S rRNA component and 21 proteins. In the eukaryotic equivalent, an 18 S rRNA component and about 33 proteins are found. In between the two subunits, a tunnel for mRNA was found. The large subunit is dome-shaped and has), site A (acceptor or aminoacyl site), enzyme peptidyl transferase and a binding site for tRNA near A site.
l The small subunit is oblate ellipsoid and has a point for recognition of mRNA and binding area for initiation factors. The small subunit fits over the large one like a cap but leaves a tunnel for mRNA. Along with the larger subunit, the small subunit from a P-site (peptidyl transfer or donor site). There are binding sites for initiation factors, elongation factors, translocase, GTP-ase, etc. The attached ribosomes are connected to membranes of endoplasmic reticulum in the region of their larger subunit by means of two types of glycoproteins, ribophorins I and II.
l The groove of the larger subunit also communicates with the channel of the endoplasmic reticulum. plays an important role in holding two subunits together (association) and also in maintaining the structure of two subunits. Soon after the completion of protein synthesis, the two subunits separate (dissociation). For this, an initiation factor (in prokaryotes, in eukaryotes) becomes attached to smaller subunits.
l The function of the ribosome is to hold in position the mRNA, tRNA and the associated enzymes controlling the process until a peptide bond forms between the adjacent amino acids.
Raw Materials for Protein Synthesis
l Amino acids, mRNA, tRNA and aminoacyl tRNA synthetase, etc. are the raw materials.
Process of Protein Synthesis
l Formation of RNA over DNA template is called transcription.
Protein synthesis consists of two main events, transcription and translation.
l It is a heterocatalytic function of DNA which involves the transfer of coded information from DNA to RNA through the synthesis of RNA over the template of DNA. Transcription is also defined as the process of copying genetic information from one strand of the DNA into RNA. Here also, the principle of complementarily governs the process of transcription, except the adenosine, now forms base pair with uracil instead of thymine.
l Transcription (written across) involves rewriting the genetic message coded in DNA into an RNA molecule. Transcription occurs in the nucleus during the and phases of the cell cycle. The mechanism of RNA synthesis was worked out in the late 1950s by American investigators Jerard Hurwitz, Samuel B. Weiss and Audrey Stevens through independent in vitro experiments. As DNA contains all the hereditary information, it is called a master copy of genetic information. It replicates its carbon copies when new master copies are required. Normally it forms working copies in the form of mRNA.
l Transcription requires a DNA template, activated precursors, a divalent metal ion, and RNA polymerase. Both strands of DNA do not transcribe RNA but only one of them called master strand or antisense strand, does. The segment of DNA involved in transcription is called cistron, the functional unit of the gene.
l Due to the following two reasons, both strands of DNA are not copied during transcription :
Ä First, if both strands act as a template, they would code for RNA molecules with different sequences, and in turn, if they code for proteins, the sequence of amino acids in the protein would be different. Hence one segment of the DNA would be coding for two different proteins, and this would complicate the genetic information transfer machinery.
Ä Second, the two RNA molecules, if produced simultaneously, would be complementary to each other, hence would form a double-stranded RNA. This would prevent RNA from being translated into protein and the exercise of transcription would become a futile one.
l A transcribing segment or transcriptional unit in DNA is defined primarily by the three regions in the DNA: a promoter, the structural gene and a terminator. There is a convention in defining the two strands of the DNA in the structural gene of a transcription unit. Since the two strands have opposite polarity and the DNA – dependent RNA polymerase also catalyse the polymerisation in only one direction, that is, the strand that has the polarity acts as a template, and is also referred to as template strand. The other strand, which has the polarity and the sequence same as RNA (except T at the location of U), is displaced during transcription. Strangely, this strand, which does not code for anything, is referred to as coding strand. All the reference points while defining a transcription unit are made with the coding strand.
l The promoter and terminator flank the structural gene in a transcription unit. The promoter, where initiation of transcription begins, is said to be located towards 5’ end (upstream) of the coding strand of the structural gene. It is a DNA sequence that provides a binding site for RNA polymerase, and it is the presence of a promoter in a transcription unit that also defines the template and coding strands. By switching its position with a terminator, the definition of coding and template strands could be reversed. The terminator is located towards 3’- end (downstream) of the coding strand and it usually defines the end of the process of transcription. There are additional regulatory sequences that may be presented further upstream and downstream to the promoter.
The enzyme required for transcription is RNA polymerase (Chamberlain and Berg, 1962; isolated from E.coli). The entire enzyme (holozyme) consists of a core enzyme and a sigma factor. The core enzyme consists of five polypeptide chains.
The holozyme has a molecular weight 4.5x. , help RNA polymerase (RNAP) in the unwinding of DNA molecules for transcription. and polypeptide also provides a catalytic basis and an active site for transcription.
l Sigmafactor recognizes the start signal or promoter region of DNA. It plays a regulatory function involving the initiation of transcription.
l Prokaryotes have only one type of RNA polymerase which synthesises all types of RNA. Eukaryotes have three types of RNA polymerases: I for rRNA (28S, 18 S and 5.8 S), II for mRNA and III for rRNAs, 5S rRNA and some Sn RNAs. Thus, there is a clear-cut division of labour. A specific protein called rho factor () is required for termination of transcription.
l Actinomycin D prevents transcription.
Steps in Transcription
Activation of Ribonucleotides
l The four types of ribonucleoside monopo-phosphates (or Ribonucleotides) – adenosine monophosphate (AMP), guanosine monophosphate (GMP), uridine monophosphate (UMP) and cytidine monophosphate (CMP) occur freely in the nucleoplasm. With the help of enzyme phosphorylase, energy and, these ribonucleoside monophosphates are changed into triphosphates – ATP, GTP, UTP and CTP, respectively.