So, HOW ARE PROTEINS MADE? PROTEIN SYNTHESIS Proteins are responsible for controlling many different processes in our bodies. As enzymes they break things down, put things together, catalyze chemical reactions, they make pigments, they form antigens and antibodies, give structure to cells and membranes, transport substances in cells and across membranes, and they perform hundreds of other functions. So, HOW ARE PROTEINS MADE?

PROTEIN SYNTHESIS Proteins are made of building blocks called AMINO ACIDS. There are 20 different amino acids. A protein can have as few as 8 aa’s or as many as 50 000 aa’s. Each protein is made by ONE GENE.

PROTEIN SYNTHESIS How does DNA, which has only 4 bases (A, T, C, and G) code for 20 different amino acids? A single base cannot code for an amino acid itself (4 bases = 4 amino acids) In pairs? Only 16 possibilities (42) AA, AT, AC, AG, TA, TT, TC, TG, CA, CT, CC, CG, GA, GT, GC, GG In triplets? 64 possibilities (43) 3 bases of DNA code for one amino acid on proteins. This is called a DNA CODON

TRANSCRIPTION DNA cannot leave the nucleus and proteins are made outside the nucleus. First the DNA must make a copy to messenger RNA (mRNA). RNA is slightly different than DNA: DNA RNA Sugar is Bases: DEOXYRIBOSE RIBOSE C, G, A, URACIL (U replace T) C, G, A, T Double stranded Single stranded

TRANSCRIPTION TRANSCRIPTION is a lot like DNA replication - the DNA unzips - but only the leading strand of DNA is copied and not as another DNA strand but as an mRNA strand. STEPS: 1) INITIATION – RNA polymerase binds to a promotor on the DNA telling it where to start transcribing. DNA is unwound.

TRANSCRIPTION STEPS: 2) RNA nucleotides are added with C matching with G and A now matching with U. This is done by an RNA POLYMERASE enzyme and is called ELONGATION.

TRANSCRIPTION STEPS: 3) Once a piece of RNA has been made, it disconnects, leaves the nucleus, and goes to the ribosomes to code for proteins.

TRANSCRIPTION SO IF A STRAND OF DNA HAS THE SEQUENCE: DNA = T A C C C G A A T C G G G T C C G A T T A C A G A T C mRNA = A U G G G C U U A G C C C A G G C U A A U G U C U A G THE STRAND OF COMPLIMENTARY mRNA WILL BE:

TRANSCRIPTION The codons (3 nucleotides) in mRNA can code for a specific amino acid, can act as INITIATORS (START codons), or TERMINATORS (STOP codons) or they can code for the same amino acid as another codon (ie. leucine has 6 different codons). This is called REDUNDANCY. TRANSCRIPTION ANIMATION

TRANSLATION The newly transcribed mRNA molecule can now detach and diffuse out of the nucleus to the cytoplasm. Once the mRNA has left the nucleus, the following steps occur: 1) mRNA binds to a RIBOSOME starting at an INITIATOR codon

TRANSLATION 2) transfer RNA (tRNA) brings specific amino acids to the ribosome. - specialized RNA called tRNA has two parts: one end binds to a specific amino acid. The other end has 3 nucleotides which bind to the codon on the mRNA. This end is the ANTICODON.

TRANSLATION 3) as the tRNA brings in an amino acid, everything shifts over one codon in the ribosome. The next tRNA brings in the next amino acid which is connected to the first with a PEPTIDE BOND and everything shifts.

TRANSLATION 4) amino acids are continually added until a TERMINATOR codon is reached on the mRNA which signals the ribosome to release the mRNA and the protein.

TRANSLATION One mRNA can be translated many times, even in a consecutive manner to produce many protein copies at once.

TRANSLATION DNA = T A C C C G A A T C G G G T C C G A T T A C A G A T C mRNA = A U G G G C U U A G C C C A G G C U A A U G U C U A G tRNA = U A C C C G A A U C G G G U C C G A U U A C A G A U C A.As = MET – GLY – LEU – ALA – GLU – ALA –ASP – VAL STOP

TRANSLATION Now YOU do it: DNA = T A C A G C T A G A A A C T G A C A A T T mRNA = A U G U C G A U C U U U G A C U G U U A A tRNA = U A C A G C U A G A A A C U G A C A A U U A.As = MET – SER – ISO – PHE – ASP – CYS STOP

TRANSLATION Now YOU do it, IN REVERSE: A.As = METH – TRYPTOPHAN– HISTIDINE- LYSINE- TYROSINE STOP mRNA = A U G U G G C A __ A A __ U A __ U A G DNA = T A C A C C G T __ T T __ A T __ A T C COMPLEMENTARY DNA = A T G T G G C A __ A A __ T A __ T A G TRANSLATION CLIP

What if you get asked about DNA when given an amino acid sequence?

What if you get asked about tRNA anticodons when given DNA code?

What if …….what else could you be asked….

Protein synthesis animations PBS website University of Utah website NCC Transcription animation NCC - Translation animation Lewis port website DNA LEARNING CENTER

F. TRANSLATION TO PROTEIN With the sequencing of the human genome, molecular biologists have isolated the genes for many important genes and their resulting proteins. While most research centers on finding the genes and proteins to help cure disease (ie. Osteoporosis, cancer, diabetes, etc.), a fringe group lead by Dr. Dickmeister, focuses their research on isolating genes for aesthetic human traits (ie. Smooth skin, hairy chest, dimples, and eye colour). It so happens that this last discovery proved to be helpful in profiling suspects using DNA evidence. One isolated gene was found to produce a protein causing blue eyes while the normal form (allele) of the gene caused brown eyes. It was discovered that the mutant blue-eye colour was the result of any mutation in the DNA causing three of the SAME amino acid to be produced side by side in the polypeptide sequence. Amazingly, it did not matter which amino acid was tripled up or where in the protein this happened – the blue eye colour would result when three of the same amino acids were side by side. Using this discovery, forensic scientists took a sample of blood from the knife, amplified the DNA, cut the DNA at known restriction sites to isolate the eye colour gene, then sequenced the DNA to elucidate the resulting polypeptide chain.

PROCEDURE: - TRANSCRIBE the segment of DNA below into mRNA. Remember: only the leading strand is transcribed. On this piece of DNA the TOP strand is the leading strand. - TRANSLATE the mRNA segment into amino acids. Remember: You must start translation at the proper START codon. If you start at the wrong place, you will produce the wrong protein. Translate the WHOLE polypeptide until you reach a STOP codon. - Determine the eye colour of the killer – if the polypeptide has three of the same amino acid in a row, the pigment produced is blue; if not, the pigment is brown. DNA SEGMENT: GGTACGGGGGTTTGCTCCAACATCGCCTGC TTCTAT CAAGTTCGACCACTTACT CCATGCCCCCAAACGAGGTTGTAGCGGACGAAGATAGTTCAAGCTGGTGAATGA

The eye colour resulting from the above polypeptide will be: ANSWER: DNA SEGMENT: GGTACGGGGGTTTGCTCCAACATCGCCTGCTTCTATCAAGTTCGACCACTTACT CCATGCCCCCAAACGAGGTTGTAGCGGACGAAGATAGTTCAAGCTGGTGAATGA mRNA SEGMENT: CCAUGCCCCCAAACGAGGUUGUAGCGGACGAAGAUAGUUCAAGCUGGUGAAUGA PROTEIN: MET(start)-PRO-PRO-ASP-GLU-VAL-VAL-ALA-ASP-GLU-ASP-SER-SER-SER-TRP-STOP EYE COLOUR: The eye colour resulting from the above polypeptide will be: THE KILLER OF GRAMPA JIM MUST BE: NOTE: The information contained in this activity is fictitious. Any similarities to actual people, places, or events is purely coincidental. DNA segments, genes, genetic traits, and proteins may have been fabricated to be representative of actual genes, traits, and proteins. No one was hurt in the making of this assignment. BLUE LORI – BLUE EYES, DAUGHTER TO NINA AND DENNY, COULD BE O- AND HAS FLAT FEET.