Guanazole is a cytostatic triazole derivative. This molecule belongs to a group of highly explosive materials, and they are vulnerable to heat, impact and friction.

Guanazole can be used in many applications, such as Chemistry and Medicine, among others. It has anti-tumor activity, by inhibition of DNA synthesis. This process occurs because this molecule inhibits ribonucleotide reductase, enzyme intervenient in the deoxyribonucleotides synthesis.

                                                  
                                                Estrutura  quimica da molécula de Guanazole

Plectonemic Structure

Plectonemic is any tertiary structure in a polymeric molecule with supercoil strands in a regular and simple form, such as supercoiled DNA.

This type of supercoiling – form shown by DNA in vivo – doesn’t produce sufficient compaction to package DNA in the cell.

In the origin of plectonemic structures there are two interwound helical filaments whose geometry is characterized by a superhelical filament angle and radius. Due to this, the superhelical angle and the twisting moment in the filaments control the action of some enzymes like topoisomerases, RNA polymerase, helycases and DNA polymerases.

Another type of supercoiling is the solenoidal form which is shown in underwound DNA. Both structures are forms of negative supercoiling and are interconvertible, though they’re two different structures.

The main advantage of plectonemic structure is it’s stability in solution, although solenoid forms can be stabilized when present in high levels in tumor cells.

Fig.1 A plectonemic helix with radius r, pitch p and opening angle a.

Image source:
Job Ubbink, Theo Odjik, Electrostatic-Undulating Theory of  Plectonemically Supercoiled DNA, Biophysical Journal, Volume 76, Issue 5 (May 1999), pp. 2502-2519, Faculty of Chemical Engineering and Materials, Delft University of Technology, 2600 GA Delft, the Netherlands

Cell senescence

Cell senescence is observed when cells stop dividing. This happens when telomeres, which protect the end of the chromosome, become progressively shorter reaching the Hayflick limit (limit to cell division capacity), meaning that human cells can only divide a finite number of times. In Cell senescence includes progressive and irreversible loss of cellular functions leading to the death of some cells.

Whenever a cell division occurs, the telomere loose length and become shorter. When they reach a minimum size, chromosome duplication ceases to occur, disabling cell division.

During this process, beta-galactosidase (enzyme responsible for hydrolysis of lactose in galactose and glucose) is detected in cell’s lysosomes, indicating senescent cells.

This process is not yet fully known, raising some doubts, which leads to the emergence of several theories. Studies show that cellular senescence may be a manifestation of loss of telomerase activity. This ribonucleoprotein is an enzyme that adds DNA sequence repeats in telomere region and thereby restores the ability of cell multiplication and retards the aging of tissues. During development, the telomerase function decline and telomeres become shorter. 

Cohesin

·     Cohesines are a member of a big protein family called SMC (Structural Maintenance of Chromosomes), who have a very important role in structural and functional organization of chromosomes.

·      SMC proteins have a crucial role in chromosome segregation and DNA repair. These proteins are interesting because of, among other aspects, their unique structure. They are dimers formed by two long coiled-coil motifs (a rod like structural shape that is formed by two long α-helices twisted around each other) connected by a non-helical sequence. An ATPase domain is created by folding a SMC monomer back to itself. This folding also creates a hinge domain at the other end. In this hinge domain, two monomers associate with each other, creating a V-shaped molecule. There are many possible structures for the SMC to possess, thanks to their high flexibility of dimers conformation.

·      Cohesines are composed by four subunits: Scc1, Scc3, Smc1 and Smc3. These last two subunits are members of SMC protein family, having therefore, ATPase domains in one end each. The two ATP domains are able to bind in the presence of ATP, thus resulting a ring-shaped structure. Scc1 and Scc3 (Spirochetal Coiled-Coil) are responsible for this binding and are also responsible of maintaining and stabilizing it. Scc1 also plays and important role in chromosome replication as it controls the separation of sister-chromatids. The cohesin ring structure is important as it keeps the sister-chromatids together during metaphase, ensuring that they travel to opposite polls of the cell (both in mitosis and meiosis). The structure facilitates as well, the spindle attachment onto chromossomes (process responsible for chromosome segregation to cell-daughters during cell division) and DNA damage repair.

Fig1: Cohesin structure

The following link is a short animation that explains the role of cohesin in cell division:

http://highered.mcgraw-hill.com/sites/9834092339/student_view0/chapter10/the_function_of_cohesin.html

Apoptosis

Apoptosis is a mechanism of programmed cell death involved in the regulation of tissue maintenance and development of organisms. It is a defense mechanism to remove excess or dangerous cells.

In this process, the chromatin condenses, cells individualize and many organelles remain intact and metabolically active for a long period. The nuclear and plasma membranes are destroyed and the nucleus is then broken up into fragments that are surrounded by the cytoplasmic membrane, producing apoptotic bodies that are degraded by macrophages. This mechanism is very distinct of necrosis, because it has no inflammatory reaction.

There are extrinsic and intrinsic apoptosis. The extrinsic pathway is triggered by the ligation of extracellular apoptotic signals to receptors in plasma membrane that lead to activation of proteins caspases in a sequence called cascade that activate other caspases or the release of cytochrome c  by the mitochondria. Intracellular apoptotic signals activate the mitochondria to release cytochrome c that indirectly activate many caspases. These proteins activate endonucleases and proteases that degrade DNA and proteins, leading to morphological changes of cells, formation of apoptotic bodies and, ultimately, cell death.

Apoptosis is a reversible process, because the blocking of apoptotic genes enhance cell survival when they are subjected to weak apoptotic signals, mutations in killer genes allow the survival of programmed cells and elimination or inhibition of macrophages result in the survival of cells.

Replicatio Factor C (RFC)

The replication factor C (Replication factor C - RFC) is a complex-key indirect involved in recruitment of the polymerase to near the replication fork and recruitment and correct positioning of complex PCNA. Composed of five subunits very conserved in eukaryotes, the complex RFC presents specific affinity for the primer-template region. The binding to the DNA molecule is ATP-dependent.

 

 

 

Fig1-Eukaryotic DNA replication: Step 1: primer synthesis by DNA polymerase - alpha (Pol-alpha); step 2: replication factor C (RFC) displacement of DNA polymerase and recruitment of proliferating cell nuclear antigen (PCNA); step 3: elongation by the newly recruited DNA polymerase-delta  holoenzyme (Pol-delta); step 4: strand displacement by Pol-delta ; step 5: cutting of the 5' displaced flap by flap endonuclease 1 (Fen1); and step 6: sealing by DNA ligase I (LigI).

 

Adriana Lima
Alexandra Meira
Bruno Oliveira

 

γ-complex

For E. coli, γ-complex is a component from the enzyme DNA polymerase III, made up of five subunits and is known by interacting with the β subunit (sliding clamp).

It’s necessary for the regulation of the attachment and removal of the DNA polymerase III from the DNA template. This function is essential for the process of attachment and detachment of the enzyme during lagging strand replication, requiring ATP. This phenomenon occurs continuously from the beginning to the end of the Okazaki fragments.

DNA Polymerase III - γ-complex