Question

RNA合成的过程?

Answer 1

RNA聚合酶进入DNA非编码区的酶切位点，解旋DNA使成为单链，核糖核苷酸由碱基互补配对法则形成RNA链（信使RNA）。

RNA的合成即DNA的转录：
DNA复制最重要的特征是半保留复制，即DNA复制时，DNA双链中的互补碱基之间的氢键断裂，解为两条单链，各以一条单链为模板，按碱基互补原则合成新的互补链，新合成的两个DNA分子和亲代DNA分子是完全一致。

在子代DNA分子中一条单链来自亲代，另一条是新合成的，这种复制方式称半保留复制(semiconservative replication)。遗传信息按这种方式忠实地从亲代传给子代。
目前有关复制的知识主要来自于原核生物实验，所以主要以原核生物来叙述。

Answer 2

遗传信息的表达是通过核酸链称为核糖核酸含量。
他们是三种类型的RNA的不同，在规模，功能和定位：
信使RNA （ mRNA的表达）是一个载体的遗传信息，复制的基因序列作为模板蛋白建设。
核糖体RNA （ S rRNA基因） ，并转移核糖核酸（ tRNA分子） （有时候也被称为不溶性和可溶性的RNA ）的结构核糖核酸酸，其中支持mRNA的表达成蛋白质。

结构

RNA的是polynucleotides链不同于那些DNA后，由核糖糖不是脱氧核糖和尿嘧啶碱基（ u ）的，而非thymines （ t ）的。

羟基功能2 '核糖大大影响性能的RNA 。尤其这使得更多大专互动往往破坏' 3 ' phosphodiester债券和防止RNA的采用抗体双螺旋构象。

但RNA的是单分子往往倍于自己的基地配对，从而形成结构所谓夏萍回路。因此，除mRNAs的陈列，平稳的线性结构， trnas和rrnas采取指定高等教育结构协会与蛋白质。

化学结构的核糖核酸
转录

在这一过程中的DNA转化为互补性的RNA （核糖核酸）的方向进行，是所谓的转录。它涉及到一个强大的酶法复杂所谓RNA聚合酶holoenzyme 。这种酶unravels和unzips的DNA螺旋结构，新兵RNA的核苷酸，火柴等，他们是由基地，以配对的DNA基因序列。

转录是相当类似，在原核生物和真核生物。其中的差异是，真核细胞具有三种不同类型的RNA聚合酶（ Ⅰ ， Ⅱ ， Ⅲ ） ，而不是一个在原核生物。每类真核生物RNA聚合酶负责合成一类的RNA （油料，我rrnas ，波兰第二的mRNA和油料三trnas和5 rrnas ） 。

转录是classicaly描述三个不同的步骤：起始，延伸和终止。

起爆发生时， RNA聚合酶holoenzyme绑定在一个特殊序列的DNA称为启动。启动子构成的一致序列含有特定字符串像塔塔（ pribnow盒）和caat （真核生物） 。

增设小蛋白，因子西格玛，重视把聚合酶，并稳定下来后，锁定对DNA链被转录。随后，聚合酶割裂了双链DNA ，形成了一个泡沫，让第一核苷三磷酸配对与互补DNA的核苷酸。

伸长的RNA链涉及连续添加核苷酸5 ' 3 '的方向。

终止发生时，停止信号，表明年底的基因是遇到了。终止信号通常是一种气相色谱-富回文形成了当地的茎环结构中的RNA ，然后由一个寡聚一个地区。这一顺序打乱了基地配对的新合成的RNA与DNA模板，迫使RNA和聚合酶跌交。有时终止还牵涉到一个特定的蛋白质（ rho蛋白） 。
转录

在原核生物细胞中，转录发生在细胞质中。当转录完成后， RNA的是马上准备用于翻译。翻译甚至可以开始在转录从而使典型的法规进程。

相比之下，真核生物转录发生在细胞核。该RNA的小学誊本，有时也被称为异种核核糖核酸（ hnrna ）往往修饰，在核出口前向细胞质中。

尤其是，真核基因进行了广泛的修改，以增加其稳定性，并成为具有生物活性。

因此， 5 '端的mRNA是盖一个7甲基（ 7mgtp ）之后不久开始。独特的5 ' -5 '三联动，形成增加m RNA的稳定性可以保障从e xonucleases。但同时也带来一个公认的信号蛋白参与随后的剪接过程中，也是在翻译过程中。

信使RNA的

Answer 3

Genetic Information is expressed through Nucleic Acids chains called RiboNucleic Acids.
They are three types of RNAs differing in size, function and localization:
Messenger RNA (mRNA) is a carrier of genetic information, a copy of a gene sequence acting as a template for protein construction.
Ribosomal RNA (rRNA) and Transfer RNA (tRNA) (also sometimes referred to as insoluble and soluble RNAs) are structural ribonucleic acids wich support the expression of mRNA into protein.

Structures

RNAs are polynucleotides chains wich differ from those of DNAs by having ribose sugar instead of deoxyribose and uracil bases(U) instead of thymines (T).

The hydroxyl function in 2' of ribose greatly affects the properties of RNAs. In particularly this enables more tertiary interactions wich tend to destabilize the 5'-3' phosphodiester bonds and prevent RNAs from adopting a B double helix conformation.

However RNAs are single stranded molecules that often fold on themselves by bases pairing, thus forming structures called hairpin loops. Thus, excepting mRNAs wich display smooth linear structure, tRNAs and rRNAs adopt specified tertiary structures in association with proteins.

The chemical structure of RiboNucleic Acid
Transcription

The process in wich DNA is converted into a complementary RNA (RiboNucleic Acid) strands is called Transcription. It involves a powerfull enzymatic complex called RNA polymerase holoenzyme. This enzyme unravels and unzips DNA helix, recruits RNA nucleotides and matches them by base pairing to the DNA gene sequence.

The transcription is rather similar in prokaryotes and eukaryotes. One of the differences is that eukariotic cells possess three different types of RNA polymerases (I, II, III), instead of one in prokaryotes. Each type of eukaryotic RNA polymerase is responsible for the synthesis of a class of RNAs (pol I for rRNAs, pol II for mRNAs and pol III for tRNAs and 5S rRNAs).

Transcription is classicaly described in three distinct steps: initiation, elongation and termination.

Initiation occurs when the RNA polymerase holoenzyme binds at a special sequence in DNA called a promoter. The promoter consists of consensus sequences containing specific strings like TATA (Pribnow box) and CAAT (in eukaryotes).

An additional small protein, the factor sigma, attaches to the polymerase and stabilises it, locking it on the DNA strand to be transcripted. Then, the polymerase separates the double stranded DNA to form a bubble allowing the first nucleoside triphosphate to pair with the complementary DNA nucleotide .

Elongation of the RNA chain involves successive addition of nucleotides in the 5' to 3' direction.

Termination occurs when a Stop signals indicating the end of the gene is encountered. The termination signal is generaly a GC-rich palindrome forming a local stem-loop structure in the RNA , followed by an oligo A region. This sequence disrupts the base pairing of newly synthesized RNA with the DNA template, forcing the RNA and the polymerase to fall off. Sometimes termination also involves a specific protein (Rho protein).
Transcription

In prokaryotes cells, transcription takes place in cytoplasm. When transcription is completed, RNAs are immediatly ready for use in translation. Translation can even begin during transcription thus allowing typical regulations process.

In contrast, eukaryotic transcription takes place in nucleus. The RNAs primary transcripts, sometimes called Heterogenous Nuclear RNA (hnRNA) are often modified in the nucleus before export to the cytoplasm.

In particular, eukaryotic mRNAs undergo extensive modifications to increase their stability and become biologically active.

Thus, the 5' end of mRNAs is capping with a 7-methylguanosine (7mGTP) shortly after initiation. The unique 5' - 5' triphosphate linkage formed increase mRNA stability by affording protection from exonucleases. It also brings a recognizable signal for proteins involved in subsequent splicing process and also during translation.

Messenger RNAs are also polyadenylated at the 3' end. Just before termination a specific sequence, AAUAAA, is recognized by a polyadenylate polymerase. The primary transcript is cleaved approximately 20 bases downstream and a string of 20 - 250 Adenines termed poly-A tail is added to the 3' end.
Eukaryotic posttranscriptional process

Since a primary transcript is a mirror copy (negatif) of all the gene sequence it includes also intronic non-conding sequences. Therefore, a post-transcriptional modification of major importance consist in introns removal in a process called RNA splicing.
The mechanism involves formation of a loop, called a lariat, in a process directed by small nuclear ribonucleoproteins (snRNPs). The complex mRNA-snRNPs is called a spliceosome.

Some proteins are often attached to exported mRNAs forming ribonucleoprotein particles (mRNP). These mRNPs are supposed to help in transport through the nuclear pores and also in binding to ribosomes.

Translation

Proteins are the major structural and functional constituants of the cells. A protein exhibit a complex molecular structure formed by polypeptide chains made of basic subunits called Amino Acids.
Expression of a mRNA code into a polypeptide chain is named Translation. This complex process requires all three classes of RNAs.
mRNAs

In the mRNA code, each amino acid is designated by a triplet of nucleotides called codon. The genetic code consist of 64 different codons (4 bases: 4x4x4 possibilities for a triplet). Three triplets are Stop codons (termination codons) which stop the process of translation. The remaining 61 codons encode 20 different amino acids. Since several codons encode a same aminoacid the genetic code is thus degenerated (or redundant).

Codons - Amino Acids connection. Click for a 3D View of the twenty amino-acids

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tRNAs

Transfert RNAs (tRNA) act as adapter between nucleotides codons and amino acids. They pick up free amino acids in cytoplasm and carry them into the ribosomes where polypeptide chain is elongated.

tRNAs are polynucleotide of about 60 - 95 nucleotides long, including few specific nucleotids (dihydro-uridine, pseudo-uridine).
They exhibit a cloverleaf-like secondary structure consisting of a stem and three main loops. They also display a tertiary L-like structure, which interacts with ribosomes.

The larger loop include a specific nucleotide triplet, the anticodon, wich may bind to a complementary codon of a mRNA.
Secondary and tertiary structure of an initiator tRNA Click for a 3D view (download time :~ 1-2 minutes)

The stem ends in 3' by the sequence ...CCA, which is the attachment site for an amino acid. Each tRNA is coupled to the amino acid in accordance with its anticodon. The coupling between a given tRNA and the corresponding amino acid is catalyzed by a specific aminoacyl-tRNA synthetases.

The different tRNAs that accept a given amino acid are called isoacceptors.
Obviously, there should be as many different tRNAs as meaning codons (ie 61). In fact there is generally at most 56 different type of tRNAs in any cell. Therefore it seems that some tRNAs are able to recognize at least two of the different codons specifying a given amino acids (Wobble hypothesis).
rRNAs

The rRNAs are the major constituents of ribosomes.
Ribosomes are the cell organelles where the mRNA is read and translated into a protein sequence. A Ribosome holds the mRNA in place, matches the anti-codon of a tRNA carring appropriate amino acid, to the complementary codon of the mRNA and catalyses the peptide bonds formation.

A ribosome consists of two subunits of different size containing rRNAs arranged with specific proteins. Both rRNAs and associated proteins are slightly different in prokaryotes vs eukaryotes.

*

The larger subunit (50S/60S) countains two rRNA molecules (5S + 23S / 5S + 28S) (S sedimentation coefficient, measures the relative size). It displays two binding sites for tRNAs : the peptidyl-tRNA (P) site and the aminoacyl-tRNA (A) site.
*

The smaller subunit (30s/40S) which is made of one rRNA molecule (16S / 18S) possesses a binding site for the mRNA.

Translation steps

Translation proceeds in cytoplasm in an ordered process. It requires free amino acids, free energy, mRNA, tRNAs, Ribosomes, and several non-ribosomal protein factors (eIF in Eukaryotes and IF in some prokaryotes).

The first phase called Initiation begins with the formation of an preinitiation complex between the small ribosomal unit, a protein factor (eIF2 or IF2) and an initiator tRNA carrying a methionine (tRNAmeti).
When the complex encounter a mRNA it recognize a specific sequence (Shine-Delgarno for prokaryotes or 5'Cap for eukaryotes) and pair the initiator codon AUG to the initiator tRNA anticodon (UAG).Then the larger ribosomal subunit associates with the initiation complex, thus matching the initiator tRNA at P site.
A next tRNA carrying an other amino acid is attracted and pairs with the next codon at the A site, the first peptide bond is catalysed by a ribosomal protein (peptidyl-transferase).

During the second phase named Elongation the ribosome continues to read codons from the 5' to the 3' and amino acids are added to the C-terminal growing peptide.
During each peptide bond formation, the polypeptide attached to the tRNA in the P site is transferred to the amino group of the aminoacyl-tRNA in the A site (Transpeptidation). Then the ribosome moves to the next codon. The empty tRNA is ejected and the peptidyl-tRNA is shifted from the A site to the P site (Translocation). A new aminoacyl-tRNA is allowed to enter within the A site.
Major steps of the Translation

Termination phase arrives when a stop codon is reached. Stop codons are triplets which are not recognized by any tRNA (UAA, UAG, UGA), but by a protein releasing factor (RF1 or RF2 in prokaryotes, eRF in eukaryotes). The factor R binds to the A site and causes the release of the polypeptide chain. The inactive ribosome then releases the mRNA and dissociates its sub-units.

It should be noted that the polypeptide sequence is in total agreement with the gene code since tRNA anticodons are complementary of mRNA codons and the mRNA sequence is a mirror of the gene DNA sequence.

Several ribosomes can progress along the same mRNA strand, each making one polypeptide chain. These clusters called polysomes are free in the cytoplasm or may be binded to particular cell organelles that store proteins (rough endoplasmic reticulum).
a polysome

References

1 Advanced Organic Chemistry of Nucleic Acids; Z. Shabarova, A. Bogdanov; VCH Verlagsgesellshaft mbH Weinheim 1994.
2 Molecular Biology of The Cell Third Ed.; Bruce Alberts et al; Garland Publishing Inc 1994.

Web References

DNA from the Beginning
Cell & Molecular Biology Online
Bio Online Book
The THMCE Medical Biochemistry Page
KU Medical Center
Surf Site for CyberBiologists
Genetic Engineering Organisation

All text & graphic contents ©1999,2000, Dr Didier Collomb all rights reserved.

For other Chemical and Biochemical pages Visit
Chemis Interactive Molecular Library

Answer 4

RNA聚合酶进入DNA非编码区的酶切位点，解旋DNA使成为单链，核糖核苷酸由碱基互补配对法则形成RNA链（信使RNA）

Answer 5

RNA的合成即DNA的转录：
DNA复制最重要的特征是半保留复制，即DNA复制时，DNA双链中的互补碱基之间的氢键断裂，解为两条单链，各以一条单链为模板，按碱基互补原则合成新的互补链，新合成的两个DNA分子和亲代DNA分子是完全一致。在子代DNA分子中一条单链来自亲代，另一条是新合成的，这种复制方式称半保留复制(semiconservative replication)。遗传信息按这种方式忠实地从亲代传给子代。
目前有关复制的知识主要来自于原核生物实验，所以主要以原核生物来叙述。复制是连续的过程，为方便叙述，可人为地把它分为起始、延长和终止三个阶段。
1.复制的起始：原核生物已固定的起始点开始，同时向各个方向进行复制，复制时双链分开成双股，新链沿张开的模板成Y字形的复制叉（replication fork）以便作为DNA合成的模板。复制起始点的辨认需多种蛋白因子，不同生物所需要的因子不同。大肠杆菌的复制起始点由DnaA蛋白识别，DnaB蛋白（解螺旋酶）在DnaC蛋白的协同下，使双链解开足够用于复制的长度，并且逐步置换出DnaA蛋白，形成引发前体复合物。DNA TopoⅡ促进复制叉的不断解链。双链解开后，SSB结合到开放的单链上，起到稳定和保护单链模板的作用，引物酶在入进来，组成引发体（primosome），包括双链DnaB、DnaC、解螺旋酶、DnaG即引物酶和DNA起始复合区域）。然后，引物酶5’→3’方向合成RNA引物，其3’-OH成为进一步合成DNA的起点。由DNA-pol Ⅲ的β亚基辨认引物，在DNA-pol Ⅲ催化下将第一个脱氧核苷酸加到引物的3’-OH上，新DNA链的合成即已开始。
2.DNA链的延长 ：在DNA-pol Ⅲ催化下自引物的3’-OH开始，沿5’→3’方向逐个地加入脱氧核苷酸，使DNA链得以延长。DNA聚合酶以3’→5’方向模板链为模板时，随着复制叉移动方向，可以连续合成前导链（leading strand）；以5’→3’方向模板链为模板合成的互补链也是沿着5’→3’方向延伸，但与复制叉的前进方向相反，只能倒着合成冈崎片段（Okaziki fragment），合成冈崎片段时，当DNA链延长到下一个引物前方时，在RNA酶和DNA-polⅠ的作用下，切除引物，并继续延长DNA链，填满切除引物后形成的空隙（gap），最后由 DNA连接酶通过生成磷酸二酯键将两个片段连接起来，封闭缺口（nick）。
3.复制的终止：复制的终止与DNA分子的形状有关。对线性DNA，当复制叉到达分子末端时，复制即终止。一般说来，DNA链复制的终止不需要特定的信号。对于环状DNA分子，两个复制叉或在一个特定部位相遇，即一个复制叉在此处停止，“等待”另一个移动较慢或需移动较长距离的复制叉，这意味着有一个特异的终止信号。