Chapter 27 Protein Metabolism 1. A brief history of understanding protein metabolism? 2. The studies leading to the deciphering of the genetic codes 3. The pathway leading to the synthesis of a functional protein; 4. Current understanding on protein targeting and degradation
Chapter 27 Protein Metabolism 1. A brief history of understanding protein metabolism; 2. The studies leading to the deciphering of the genetic codes; 3. The pathway leading to the synthesis of a functional protein; 4. Current understanding on protein targeting and degradation
1. Translation(protein synthesis) necessitates the coordinated interplay of about 300 macromolecules in the cells The most complex of all biosynthetic path ways 60 to 90 macromolecules for making up the protein synthesizing machine ribosomes Over 20 enzymes for activating the amino acids Over 10 auxiliary proteins for the initiation, elongation and termination of the polypeptide chains Account for up to 90% of the chemical energy used by a cell for all biosynthetic reactions
1. Translation (protein synthesis) necessitates the coordinated interplay of about 300 macromolecules in the cells • The most complex of all biosynthetic pathways. • 60 to 90 macromolecules for making up the proteinsynthesizing machine ribosomes • Over 20 enzymes for activating the amino acids. • Over 10 auxiliary proteins for the initiation, elongation and termination of the polypeptide chains. • Account for up to 90% of the chemical energy used by a cell for all biosynthetic reactions
The molecules used for translation account for more than 35%o of the cells dry weight However, proteins are synthesized with very high efficiency: a complete polypeptide chain of 100 residues is synthesized in about 5 seconds in an E coli cells at 37oc
• The molecules used for translation account for more than 35% of the cell’s dry weight. • However, proteins are synthesized with very high efficiency: a complete polypeptide chain of 100 residues is synthesized in about 5 seconds in an E.coli cells at 37oC
2. The molecular mechanism of protein synthesis was mainly revealed during the 2nd half of the 20th century Ribonucleoprotein particles(were later called ribosomes) were revealed to be the site of protein synthesis in rat liver cells, using radioactively labeled amino acids and immediate subcellular fractionations(early 1950s, by zamecnik Amino acids were found to be activated by attaching to a special form of heat-stable RNa molecules(later called tRNas before being incorporated into polypeptides(1950s by hoagland and zamecnik)
2. The molecular mechanism of protein synthesis was mainly revealed during the 2nd half of the 20th century • Ribonucleoprotein particles (were later called ribosomes) were revealed to be the site of protein synthesis in rat liver cells, using radioactively labeled amino acids and immediate subcellular fractionations (early 1950s, by Zamecnik). • Amino acids were found to be activated by attaching to a special form of heat-stable RNA molecules (later called tRNAs) before being incorporated into polypeptides (1950s, by Hoagland and Zamecnik)
Each tRNa molecule was found to function as an adapter (originally hypothesized by francis Crick) carrying a specific amino acid with one site and recognizing a specific site on a template with another site The concept of messenger RNa(mRNA) was boldly formulated by Jacob and Monod in 1961: a short-lived RNA Should serve as the information carrier between gene and protein(to explain the quick induction of proteins in E coli) This bold hypothesis was quickly confirmed by studies of E coli cells infected by T2 phages
• Each tRNA molecule was found to function as an adapter (originally hypothesized by Francis Crick), carrying a specific amino acid with one site and recognizing a specific site on a template with another site. • The concept of messenger RNA (mRNA) was boldly formulated by Jacob and Monod in 1961: a short-lived RNA should serve as the information carrier between gene and protein (to explain the quick induction of proteins in E.coli). • This bold hypothesis was quickly confirmed by studies of E.coli cells infected by T2 phages
Ribosomes Ribosomes were revealed to be the site of protein synthesis in early 1950s(pulse labeling with radioactive amino acids and subcellular fractionations)
Ribosomes were revealed to be the site of protein synthesis in early 1950s (pulse labeling with radioactive amino acids and subcellular fractionations)
Crick's Amino acid H molecule adapter O2C-C-NH3 hypothesis R Amino acid bindi ng site Hydroger . Adapter(tRNA bonds C UIA UGA C UA G UC GG mrNA Nucleotide triplet coding for an amino acid
Crick’s adapter hypothesis Hydrogen bonds
3 Amino acids in a polypeptide chain were found to be coded by groups of three nucleotides in a mrNa Simple calculation indicated that three or more bases are probably needed to specify one amino acid Genetic studies of insertion deletion and substitution mutants showed codons for amino acids are triplet of nucleotides; codons do not overlap and there is no punctuation between codons for successive amino acid residues The amino acid sequence of a polypeptide is defined by a linear sequence of contiguous codons: the first codon establishs a reading frame
3. Amino acids in a polypeptide chain were found to be coded by groups of three nucleotides in a mRNA • Simple calculation indicated that three or more bases are probably needed to specify one amino acid. • Genetic studies of insertion, deletion, and substitution mutants showed codons for amino acids are triplet of nucleotides; codons do not overlap and there is no punctuation between codons for successive amino acid residues. • The amino acid sequence of a polypeptide is defined by a linear sequence of contiguous codons: the first codon establishs a reading frame
mRNA 5'--G U C CUA CG G AU---3 (+) Insertion-GUAG C CUC AC G GA U Altered amino acid sequences Deletion. - UAC C UA C GG A U (平) Insertion and G U AA G CCA CIG G AU deletion Reading frame restored Genetic studies showed that genetic codons are successive triplets of nucleotides
Genetic studies showed that genetic codons are successive triplets of nucleotides Altered amino acid sequences
Nonoverlapping A U ACG AG U C code 1 2 3 Overlapping A UACGAGU C de Amino acid sequence studies of tobacco mosaic virus mutants and abnormal hemoglobins showed that alterations usually affected only one single amino acid: genetic codes are nonoverlapping
Amino acid sequence studies of tobacco mosaic virus mutants and abnormal hemoglobins showed that alterations usually affected only one single amino acid: genetic codes are nonoverlapping