Instructor: Dr. Natalia Tretyakova, Ph.D. «hyperlink "mailto:[email protected]"»    -   6-3432
 PDB reference correction and design Dr.chem., Ph.D. Aris Kaksis, Associate Prof. mailto:[email protected]
Transcription                                                        Transcription Translation
                                        Translation
 
 
 
Protein Synthesis Part 1: Take Home Message 
 Required reading: Stryer Ch. 34, p. 875-888 Ch. 33 p. 849-850
 
 1) Translation of the genetic code is dependent on three base words that correspond to a single
                                                                                                            amino acid AA (codons).
2) The mRNA message is read by tRNA through the use of a three base complement to the three 3
                                                                                                                base word (anticodon).
3) A specific amino acid AA is conjugated to a specific tRNA.
4) Amino acid AA side chain size, hydrophobicity and polarity govern the ability of tRNA synthetases
                                                   to conjugate a specific three 3 base message with a specific amino acid AA.
 
Table 1. The genetic code. for mRNA Y                                          Genetic Code 
 
1st position 2nd position 3rd position
(5' end)--> C A G (3' end)-->
Phe
Phe
Leu
Leu 		Ser
Ser
Ser
Ser 		Tyr
Tyr
STOP
STOP 		Cys
Cys
STOP SelenoCys Trp Mitochondria
Trp
C
A
G
C Leu
Leu
Leu
Leu 		Pro
Pro
Pro
Pro 		His
His
Gln
Gln 		Arg
Arg
Arg
Arg
C
A
G
A Ile
Ile
Ile
Met init 		Thr
Thr
Thr
Thr 		Asn
Asn
Lys
Lys 		Ser
Ser
Arg
Arg
C
A
G
G Val
Val
Val
Val 		Ala
Ala
Ala
Ala 		Asp
Asp
Glu
Glu 		Gly
Gly
Gly
Gly
C
A
G
 
Sets of three 3 nucleotides (codons) in an mRNA molecule are translated into amino acids AA in the course of protein synthesis according to the rules shown. The codons GG and GAG, for example, are translated into valine and glutamic acid, respectively. Note that those codons with  or C as the second 2 nucleotide tend to specify the more hydrophobic amino acids AA.
 
Structure of Transfer RNA
 

Di-hydro-uridine (DHU)                   Anti Codon                            Pseudo-uridine (Y)
 
Classes of Aminoacyl-tRNA Synthetases
 
• Class I: Arg, Cys, Gln, Glu, Ile, Leu, Met, Trp, Tyr, Val    (Generally the Larger Amino Acids)
• Class II: Ala, Asn, Asp, Gly, His , Lys, Phe, Ser, Pro, Thr   (Generally the smaller amino acids)
                                                                                      Charged tRNA -76 
 
tRNA Activation by aminoacyl tRNA synthases
 
1. Amino-acyl-AMP formation

                                                                           2. Aminoacyl transfer to the appropriate tRNA + AMP1-

 
             tRNA Recognition by Synthetases
 
• Specific recognition of the anticodon; ex. an change the anticodon for tRNATrp to
                                                                                  that for methionine and get good acylation by tRNAMet.
•Stem sequences can be crucial; ex. tRNAAla depends on GC at position 3:70 doesn't care what the codon is.
• Both the stem regions and anticodon are needed; ex. tRNA Gln.
 
tRNA Recognition by Synthetases 
 
        tRNA-Ala                                         tRNA-Phe                                     Trna- Ser

 
tRNA Recognition by Synthetases 
 
          Anticodon Recognition ­
 
tRNA Synthetase Proofreading Ile
 
         Large         Smaller                                                               Large                Smaller
Acylation Site Hydrolytic Site                                                 Acylation Site Hydrolytic Site
<-- Difference --
                                            
Correct Acylation                                                                                                                         Mis-acylation
tRNA Synthetase Proofreading Val
 
Hydrophobic      Polar                                                               Hydrophobic      Polar
Acylation Site Hydrolytic Site                                                 Acylation Site Hydrolytic Site
¬ Difference ®
                                                   
  Correct Acylation                                                                                                                  Mis-acylation
 
How do we go from mRNA to Protein?
 

Incoming tRNA
                                           Table 1. The genetic code. for mRNA Y                                                                          Genetic Code 
 
1st position 2nd position 3rd position
(5' end)--> C A G (3' end)-->
Phe
Phe
Leu
Leu 		Ser
Ser
Ser
Ser 		Tyr
Tyr
STOP
STOP 		Cys
Cys
STOP SelenoCys Trp Mitochondria
Trp
C
A
G
C Leu
Leu
Leu
Leu 		Pro
Pro
Pro
Pro 		His
His
Gln
Gln 		Arg
Arg
Arg
Arg
C
A
G
A Ile
Ile
Ile
Met init 		Thr
Thr
Thr
Thr 		Asn
Asn
Lys
Lys 		Ser
Ser
Arg
Arg
C
A
G
G Val
Val
Val
Val 		Ala
Ala
Ala
Ala 		Asp
Asp
Glu
Glu 		Gly
Gly
Gly
Gly
C
A
G
 
Sets of three 3 nucleotides (codons) in an mRNA molecule are translated into amino acids AA in the course of protein synthesis according to the rules shown. The codons GG and GAG, for example, are translated into valine and glutamic acid, respectively. Note that those codons with  or C as the second 2 nucleotide tend to specify the more hydrophobic amino acids AA.
 

 
                                              Wobble pairing rules
5’ anticodon base         3’ codon base
 C                                   G
 A                                  
                              A or G
 G                               C or  
I                             , C, or A
                                                       tRNA Anticodon-Codon Recognition 
 
Anticodon 3’ – C – G –  I – 5’               3’ – C – G –  I – 5’                   3’ – C – G –  I – 5’
Codon       5’ – C – G – – 3’               5’ – C – G – C – 3’                   5’ – C – G – A – 3’
 
Anticodon 3’ – C – G – G – 5’               3’ – C – G – G – 5’
Codon       5’ – C – G – – 3’               5’ – C – G – C – 3’
 
Anticodon 3’ – C – G – – 5’               3’ – C – G – – 5’
Codon       5’ – C – G – A – 3’               5’ – C – G – G – 3’
 
Anticodon 3’ – C – G – C – 5’               3’ – C – G – A – 5’
Codon       5’ – C – G – G – 3’               5’ – C – G – – 3’
 
tRNA Anticodon-Codon Recognition
 
              Adenosine                                Inosine                                                      Guanosine
                                                
 Anticodon 3’ – C – G – I – 5’                3’ – C – G – I – 5’                    3’ – C – G – I – 5’
Codon       5’ – C – G – C – 3’               5’ – C – G – A – 3’                   5’ – C – G – – 3’
         
              I------C  base pair                   I-----A  base pair                               I----- base pair
 
Nonsense suppression
 
   
 
Table 1. The genetic code. for mRNA Y                                                                        Genetic Code
 
1st position 2nd position 3rd position
(5' end)--> C A G (3' end)-->
Phe
Phe
Leu
Leu 		Ser
Ser
Ser
Ser 		Tyr
Tyr
STOP
STOP 		Cys
Cys
STOP SelenoCys Trp Mitochondria
Trp
C
A
G
C Leu
Leu
Leu
Leu 		Pro
Pro
Pro
Pro 		His
His
Gln
Gln 		Arg
Arg
Arg
Arg
C
A
G
A Ile
Ile
Ile
Met init 		Thr
Thr
Thr
Thr 		Asn
Asn
Lys
Lys 		Ser
Ser
Arg
Arg
C
A
G
G Val
Val
Val
Val 		Ala
Ala
Ala
Ala 		Asp
Asp
Glu
Glu 		Gly
Gly
Gly
Gly
C
A
G
 
Sets of three 3 nucleotides (codons) in an mRNA molecule are translated into amino acids AA in the course of protein synthesis according to the rules shown. The codons GG and GAG, for example, are translated into valine and glutamic acid, respectively. Note that those codons with  or C as the second 2 nucleotide tend to specify the more hydrophobic amino acids AA.
 
Protein Synthesis Part 2: Take Home Message
 1) Translation always proceeds by reading the mRNA sequence from the 5’ to the 3’ end.
 2) Amino acids are assembled in to proteins with the first amino acid having a free amino group
         —NH3+   (N-terminus) and the last having a free carboxylate—COO— (C-terminus).
  3) Synthesis of the protein is processive and contains steps;
                                                                                                  a) initiation, b) elongation and c) termination
 
 Overall Ribosome Structure:                               Translation Machine


• tRNA site, peptidyl transferase, GTPase and Exit site
  DNA        5'-ATG---GCC---GA--C-AAA--AA- 3’       Translating the Message
RNA       5' -AG--GCC---GA--C-AAA--AA--3’
Protein      N-  Met     Ala    Phe        Asp      Ser     Lys   Stop -C         Overview of Translation
 

 1) Translation of the genetic code is dependent on three base words
                                                                                                             that correspond to a single amino acid.
2) The mRNA message is read by tRNA through the use of a three base complement to
                                                                                                                                           the three 3  base word.
3) A specific amino acid is conjugated to a specific tRNA (three base word).
4) Amino acid side chain size, hydrophobicity and polarity govern the ability of tRNA synthetases to
                                                        conjugate a specific three base message with a specific amino acid AA.