SINTESIS PROTEIN Sri Puji Astuti Wahyuningsih FST UNAIR    BIODAS II Dept. Biologi FST UNAIR

ARUS INFORMASI GENETIK Informasi DNA adalah dalam bentuk urutan tertentu dari nukleotida di sepanjang untai DNA DNA diwariskan oleh organisme Menghasilkan ciri-ciri tertentu melalui sintesis protein Proses di mana DNA mensintesis protein dinamakan ekspresi gen Ada dua tahapan, yaitu transkripsi dan translasi BIODAS II Dept. Biologi FST UNAIR

RIBOSOM Adalah organel sel yang menfasilitasi terjadinya translasi (sintesis polipeptida) Figure 17.1 BIODAS II Dept. Biologi FST UNAIR

Konsep 1: GEN MENENTUKAN PROTEIN MELALUI TRANSKRIPSI DAN TRANSLASI 1909, Dokter Inggris Archibald Garrod Adalah orang pertama yang menunjukkan bahwa gen menentukan fenotip melalui enzim yang mengkatalisis reaksi kimia tertentu dalam sel 1920, Beadle dan Edward Tatum: Neurospora crassa (kapang roti) yang dimutasi dengan sinar-X Menciptakan mutan yang tidak bisa bertahan hidup pada media minimal Mengembangkan hipotesis "satu gen-satu enzim“ Fungsi gen adalah untuk menentukan produk enzim tertentu BIODAS II Dept. Biologi FST UNAIR

PRINSIP DASAR TRANSKRIPS DAN TRANSLASI Transkripsi Adalah sintesis RNA di bawah arahan DNA Menghasilkan messenger RNA (mRNA) atau RNA duta Translasi Adalah sintesis polipeptida yang terjadi di bawah arahan mRNA Terjadi dalam ribosom BIODAS II Dept. Biologi FST UNAIR

Pada Prokariota Transkripsi dan translasi terjadi bersamaan (translasi mRNA dimulai saat transkripsi masih berlangsung) Prokaryotic cell. In a cell lacking a nucleus, mRNA produced by transcription is immediately translated without additional processing. (a) TRANSLATION TRANSCRIPTION DNA mRNA Ribosome Polypeptide Figure 17.3a BIODAS II Dept. Biologi FST UNAIR

Pada Eukariota Transkrip RNA atau transkrip primer atau pre-mRNA dimodifikasi sebelum menjadi mRNA Figure 17.3b Eukaryotic cell. The nucleus provides a separate compartment for transcription. The original RNA transcript, called pre-mRNA, is processed in various ways before leaving the nucleus as mRNA. (b) TRANSCRIPTION RNA PROCESSING TRANSLATION mRNA DNA Pre-mRNA Polypeptide Ribosome Nuclear envelope BIODAS II Dept. Biologi FST UNAIR

DNA RNA protein Aliran informasi genetik adalah Dikodekan sebagai urutan triplet basa yang tidak tumpang tindih yang disebut kodon BIODAS II Dept. Biologi FST UNAIR

Selama transkripsi Gen menentukan urutan basa di sepanjang molekul mRNA Figure 17.4 DNA molecule Gene 1 Gene 2 Gene 3 DNA strand (template) TRANSCRIPTION mRNA Protein TRANSLATION Amino acid A C G T U Trp Phe Gly Ser Codon 3 5 BIODAS II Dept. Biologi FST UNAIR

KODE GENETIK Kodon dalam mRNA Diterjemahkan menjadi asam amino atau berfungsi sebagai sinyal mulai (start) dan berhenti (stop) dari translasi Figure 17.5 Second mRNA base U C A G UUU UUC UUA UUG CUU CUC CUA CUG AUU AUC AUA AUG GUU GUC GUA GUG Met or start Phe Leu lle Val UCU UCC UCA UCG CCU CCC CCA CCG ACU ACC ACA ACG GCU GCC GCA GCG Ser Pro Thr Ala UAU UAC UGU UGC Tyr Cys CAU CAC CAA CAG CGU CGC CGA CGG AAU AAC AAA AAG AGU AGC AGA AGG GAU GAC GAA GAG GGU GGC GGA GGG UGG UAA UAG Stop UGA Trp His Gln Asn Lys Asp Arg Gly First mRNA base (5 end) Third mRNA base (3 end) Glu BIODAS II Dept. Biologi FST UNAIR

Komponen Molekular Transkripsi Konsep 2: TRANSKRIPSI ADALAH SINTESIS RNA YANG DIARAHKAN DNA Komponen Molekular Transkripsi Sintesis RNA dikatalisis oleh RNA polimerase. Berfungsi untuk memisahkan untai DNA dan menghasilkan nukleotida RNA dari arah 5’  3’ Prokariota hanya mempunyai 1 jenis RNA polimerase. Eukatiota memiliki 3 jenis RNA polimerase, yang digunakan untuk sintesis mRNA adalah RNA polimerase II BIODAS II Dept. Biologi FST UNAIR

Sintesis Transkrip RNA Tahapan transkripsi Inisiasi Elongasi Terminasi Figure 17.7 Promoter Transcription unit RNA polymerase Start point 5 3 Rewound RNA transcript Completed RNA transcript Unwound DNA Template strand of DNA 1 Initiation. After RNA polymerase binds to the promoter, the DNA strands unwind, and the polymerase initiates RNA synthesis at the start point on the template strand. 2 Elongation. The polymerase moves downstream, unwinding the DNA and elongating the RNA transcript 5  3 . In the wake of transcription, the DNA strands re-form a double helix. 3 Termination. Eventually, the RNA transcript is released, and the polymerase detaches from the DNA. BIODAS II Dept. Biologi FST UNAIR

Pengikatan RNA Polimerase dan Inisiasi Transkripsi Figure 17.8 TRANSCRIPTION RNA PROCESSING TRANSLATION DNA Pre-mRNA mRNA Ribosome Polypeptide T A TATA box Start point Template DNA strand 5 3 Transcription factors Promoter RNA polymerase II Transcription factors RNA transcript Transcription initiation complex Eukaryotic promoters 1 Several transcription 2 Additional transcription 3 Promotor : sekuen DNA tempat melekatnya RNA polimerase dan tempat inisiasi untuk sintesis RNA Faktor Transkripsi Membantu RNA polimerase untuk mengenali sekuen promoter pada Eukariota Figure 17.8 BIODAS II Dept. Biologi FST UNAIR

Elongation RNA polymerase Non-template strand of DNA RNA nucleotides 3 end C A E G U T 3 5 Newly made Direction of transcription (“downstream”) Template BIODAS II Dept. Biologi FST UNAIR

Terminasi Transkripsi Elongasi Transkripsi RNA polimerase bergerak sepanjang DNA Enzim menguraikan DNA double helix, sekitar 10- 20 basa DNA dibuka dan dibaca untuk menghasilkan pasangannya, yaitu nukleotida RNA Terminasi Transkripsi RNA polimerase telah selesai membaca DNA. Telah dihasilkan seluruh untaian nukleotida RNA BIODAS II Dept. Biologi FST UNAIR

Konsep 3: SEL EUKARIOTIK MEMODIFIKASI RNA SETELAH TRANSKRIPSI Enzim di inti sel pada eukariotik Modifikasi pre-mRNA dengan cara tertentu sebelum pesan genetik dikirim ke sitoplasma BIODAS II Dept. Biologi FST UNAIR

Pengubahan ujung mRNA Setiap ujung molekul pre-mRNA dimodifikasi dengan cara tertentu Ujung 5 dimodifikasi dengan penambahan nukeotida tudung (cap) Ujung 3 dengan penambahan ekor poli-A A modified guanine nucleotide added to the 5 end 50 to 250 adenine nucleotides added to the 3 end Protein-coding segment Polyadenylation signal Poly-A tail 3 UTR Stop codon Start codon 5 Cap 5 UTR AAUAAA AAA…AAA TRANSCRIPTION RNA PROCESSING DNA Pre-mRNA mRNA TRANSLATION Ribosome Polypeptide G P 5 3 Figure 17.9 BIODAS II Dept. Biologi FST UNAIR

Split Genes dan RNA Splicing Penyambungan RNA / RNA splicing Memindahkan intron dan menggabungkan exon TRANSCRIPTION RNA PROCESSING DNA Pre-mRNA mRNA TRANSLATION Ribosome Polypeptide 5 Cap Exon Intron 1 5 30 31 104 105 146 3 Poly-A tail Introns cut out and exons spliced together Coding segment 3 UTR Figure 17.10 BIODAS II Dept. Biologi FST UNAIR

RNA transcript (pre-mRNA) Dilakukan oleh spliceosome, yaitu enzim yang berfungsi untuk memindahkan intron dan menggabungkan exon RNA transcript (pre-mRNA) Exon 1 Intron Exon 2 Other proteins Protein snRNA snRNPs Spliceosome components Cut-out intron mRNA 5 1 2 3 Figure 17.11 BIODAS II Dept. Biologi FST UNAIR

Fungsi dan Pentingnya Intron Adanya intron Memungkinkan satu gen dapat mengkode lebih dari satu jenis polipeptida BIODAS II Dept. Biologi FST UNAIR

Protein sering memiliki arsitektur modular Terdiri dari daerah struktural dan fungsional yang disebut domain Kode exon akan berbeda untuk domain yang berbeda pada protein Gene DNA Exon 1 Intron Exon 2 Exon 3 Transcription RNA processing Translation Domain 3 Domain 1 Domain 2 Polypeptide Figure 17.12 BIODAS II Dept. Biologi FST UNAIR

Komponen Molekular Translasi Konsep 4: TRANSLASI ADALAH SINTESIS POLIPEPTIDA YANG DIARAHKAN OLEH RNA Komponen Molekular Translasi Sel mentranslasi pesan mRNA menjadi protein dengan bantuan RNA transfer (tRNA) BIODAS II Dept. Biologi FST UNAIR

Konsep Dasar Translasi TRANSCRIPTION TRANSLATION DNA mRNA Ribosome Polypeptide Amino acids tRNA with amino acid attached tRNA Anticodon Trp Phe Gly A G C U Codons 5 3 Figure 17.13 BIODAS II Dept. Biologi FST UNAIR

Struktur dan Funsi RNA Transfer Molekul tRNA Tersusun dari untai RNA tunggal yang panjangnya hanya sekitar 80 nukleotida Berbentuk L A C C Figure 17.14a Two-dimensional structure. The four base-paired regions and three loops are characteristic of all tRNAs, as is the base sequence of the amino acid attachment site at the 3 end. The anticodon triplet is unique to each tRNA type. (The asterisks mark bases that have been chemically modified, a characteristic of tRNA.) (a) 3 C A G U * 5 Amino acid attachment site Hydrogen bonds Anticodon BIODAS II Dept. Biologi FST UNAIR

(b) Three-dimensional structure Symbol used in this book Amino acid attachment site Hydrogen bonds Anticodon A G 5 3 (c) Figure 17.14b BIODAS II Dept. Biologi FST UNAIR

Enzim aminoacyl-tRNA synthetase Menggabungkan masing-masing asam amino ke tRNA yang benar Amino acid Aminoacyl-tRNA synthetase (enzyme) 1 Active site binds the amino acid and ATP. P P P Adenosine ATP ATP loses two P groups and joins amino acid as AMP. 2 P Adenosine Pyrophosphate P Pi Pi Phosphates Pi tRNA 3 Appropriate tRNA covalently Bonds to amino Acid, displacing AMP. P Adenosine AMP Activated amino acid is released by the enzyme. 4 Aminoacyl tRNA (an “activated amino acid”) Figure 17.15 BIODAS II Dept. Biologi FST UNAIR

Ribosom Menfasilitasi ikatan antikodon tRNA dengan kodon mRNA selama sintesis protein TRANSCRIPTION TRANSLATION DNA mRNA Ribosome Polypeptide Exit tunnel Growing polypeptide tRNA molecules E P A Large subunit Small Computer model of functioning ribosome. This is a model of a bacterial ribosome, showing its overall shape. The eukaryotic ribosome is roughly similar. A ribosomal subunit is an aggregate of ribosomal RNA molecules and proteins. (a) 5 3 Subunit ribosomal Disusun oleh protein dan molekul RNA yang dinamakan RNA ribosomal atau rRNA Figure 17.16a BIODAS II Dept. Biologi FST UNAIR

Ribosom mempunyai 3 sisi pengikatan (binding sites) tRNA Sisi A Sisi E P site (Peptidyl-tRNA binding site) E site (Exit site) mRNA binding site A site (Aminoacyl- tRNA binding site) Large subunit Small Schematic model showing binding sites. A ribosome has an mRNA binding site and three tRNA binding sites, known as the A, P, and E sites. This schematic ribosome will appear in later diagrams. (b) E P A Figure 17.16b BIODAS II Dept. Biologi FST UNAIR

Growing polypeptide Amino end Next amino acid to be added to polypeptide chain tRNA mRNA Codons 3 5 Schematic model with mRNA and tRNA. A tRNA fits into a binding site when its anticodon base-pairs with an mRNA codon. The P site holds the tRNA attached to the growing polypeptide. The A site holds the tRNA carrying the next amino acid to be added to the polypeptide chain. Discharged tRNA leaves via the E site. (c) Figure 17.16c BIODAS II Dept. Biologi FST UNAIR

Pembentukan Polipeptida Kita dapat membagi translasi menjadi 3 tahapan Inisiasi Elongasi Terminasi BIODAS II Dept. Biologi FST UNAIR

Asosiasi Ribosom dan Inisiasi Translasi Tahap inisiasi translasi Menyatukan mRNA, tRNA yang membawa asam amino pertama dari polipeptida, dan dua subunit ribosom Large ribosomal subunit The arrival of a large ribosomal subunit completes the initiation complex. Proteins called initiation factors (not shown) are required to bring all the translation components together. GTP provides the energy for the assembly. The initiator tRNA is in the P site; the A site is available to the tRNA bearing the next amino acid. 2 Initiator tRNA mRNA mRNA binding site Small Translation initiation complex P site GDP GTP Start codon A small ribosomal subunit binds to a molecule of mRNA. In a prokaryotic cell, the mRNA binding site on this subunit recognizes a specific nucleotide sequence on the mRNA just upstream of the start codon. An initiator tRNA, with the anticodon UAC, base-pairs with the start codon, AUG. This tRNA carries the amino acid methionine (Met). 1 Met U A C G E 3 5 Figure 17.17 BIODAS II Dept. Biologi FST UNAIR

Elongasi Translasi Asam amino ditambahkan satu per satu ke asam amino sebelumnya Codon recognition. The anticodon of an incoming aminoacyl tRNA base-pairs with the complementary mRNA codon in the A site. Hydrolysis of GTP increases the accuracy and efficiency of this step. 1 Figure 17.18 Amino end of polypeptide mRNA Ribosome ready for next aminoacyl tRNA E P A GDP GTP 2 site 5 3 TRANSCRIPTION TRANSLATION DNA Ribosome Polypeptide Translocation. The ribosome translocates the tRNA in the A site to the P site. The empty tRNA in the P site is moved to the E site, where it is released. The mRNA moves along with its bound tRNAs, bringing the next codon to be translated into the A site. 3 Peptide bond formation. An rRNA molecule of the large subunit catalyzes the formation of a peptide bond between the new amino acid in the A site and the carboxyl end of the growing polypeptide in the P site. This step attaches the polypeptide to the tRNA in the A site. 2 BIODAS II Dept. Biologi FST UNAIR

Terminasi Translasi Tahap akhir translasi adalah ketika ribosom sampai kodon stop pada mRNA Figure 17.19 Release factor Free polypeptide Stop codon (UAG, UAA, or UGA) 5 3 When a ribosome reaches a stop codon on mRNA, the A site of the ribosome accepts a protein called a release factor instead of tRNA. 1 The release factor hydrolyzes the bond between the tRNA in the P site and the last amino acid of the polypeptide chain. The polypeptide is thus freed from the ribosome. 2 3 The two ribosomal subunits and the other components of the assembly dissociate. BIODAS II Dept. Biologi FST UNAIR

Poliribosom Sejumlah ribosom dapat mentranslasi satu molekul mRNA tunggal secara simultan. Hanya pada sel prokariota Figure 17.20a, b Growing polypeptides Completed polypeptide Incoming ribosomal subunits Start of mRNA (5 end) End of (3 end) Polyribosome An mRNA molecule is generally translated simultaneously by several ribosomes in clusters called polyribosomes. (a) Ribosomes This micrograph shows a large polyribosome in a prokaryotic cell (TEM). 0.1 µm (b) BIODAS II Dept. Biologi FST UNAIR

Protein Fungsional Rantai polipeptida /protein menjalani modifikasi setelah proses translasi Protein dimodifikasi - untuk membentuk molekul tiga dimensi / folding (pelipatan) - Untuk dapat ditranfer ke lokasi yang membutuhkan BIODAS II Dept. Biologi FST UNAIR

Protein dibutuhkan dalam sistem endomembran (contoh: enzim lisosim yang ada di organel lisosom) atau disekresikan (contoh: enzim pencernaan seperti amilase). Protein harus ditransport ke retikulum endoplasma (ER) Signal peptida pada ribosom binding pada signal-recognition particle (SRP). Kompleks keduanya binding pada protein reseptor SRP di ER BIODAS II Dept. Biologi FST UNAIR

Mekanisme signal untuk protein target pada ER Polypeptide synthesis begins on a free ribosome in the cytosol. 1 An SRP binds to the signal peptide, halting synthesis momentarily. 2 The SRP binds to a receptor protein in the ER membrane. This receptor is part of a protein complex (a translocation complex) that has a membrane pore and a signal-cleaving enzyme. 3 The SRP leaves, and the polypeptide resumes growing, meanwhile translocating across the membrane. (The signal peptide stays attached to the membrane.) 4 The signal- cleaving enzyme cuts off the signal peptide. 5 The rest of the completed polypeptide leaves the ribosome and folds into its final conformation. 6 Figure 17.21 Ribosome mRNA Signal peptide Signal- recognition particle (SRP) SRP receptor protein Translocation complex CYTOSOL removed ER membrane Protein ERLUMEN

Ringkasan transkripsi dan translasi pada sel eukariotik TRANSCRIPTION RNA is transcribed from a DNA template. DNA RNA polymerase transcript RNA PROCESSING In eukaryotes, the RNA transcript (pre- mRNA) is spliced and modified to produce mRNA, which moves from the nucleus to the cytoplasm. Exon Poly-A RNA transcript (pre-mRNA) Intron NUCLEUS Cap FORMATION OF INITIATION COMPLEX After leaving the nucleus, mRNA attaches to the ribosome. CYTOPLASM mRNA Growing polypeptide Ribosomal subunits Aminoacyl-tRNA synthetase Amino acid tRNA AMINO ACID ACTIVATION Each amino acid attaches to its proper tRNA with the help of a specific enzyme and ATP. Activated amino acid TRANSLATION A succession of tRNAs add their amino acids to the polypeptide chain as the mRNA is moved through the ribosome one codon at a time. (When completed, the polypeptide is released from the ribosome.) Anticodon A C U G E Ribosome 1 5 3 Codon 2 3 4 5 Figure 17.26 BIODAS II Dept. Biologi FST UNAIR

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