WHAT IS PROTEIN DATA BANK ( PDB ) ??
lA repository for 3-D biological macromolecular structure
lAll data are available to the public
lIt includes proteins, nucleic acids and viruses
lObtained by X-Ray crystallography (80%) or NMR spectroscopy (16%)
lSubmitted by biologists and biochemists from around the world
lPDB is an important resource for research in the academic, pharmaceutical, and biotechnology sectors
lExamples
–Will this molecule turns into a cancer cell?
–Can this combination of molecules cure common cold?
–How does radiation affect the RNA and DNA?
lFounded in 1971 by Brookhaven National Laboratory, New York
lFirst set of data were entered on punched cards. Then with magnetic tapes
lTransferred to the Research Collaborators for Structural Bioinformatics (RCSB) in 1998
lCurrently it holds 29,000 released structures
1ST MOLECULE : SUBTILISIN
Synonyms: Subtilisin Carlsberg, Subtilopeptidase A, Bacterial Alkaline Protease
Subtilisins belong to subtilases, a group of serine proteases that initiate the nucleophilic attack on the peptide (amide) bond through a serine residue at the active site. They are physically and chemically well-characterized enzymes. Subtilisins typically have molecular weights of about 20,000 to 45,000 dalton. They can be obtained from soil bacteria, for example, Bacillus amyloliquefaciens. Subtilisins are secreted in large amounts from many Bacillus species. Subtilisins are widely used in commercial products, for example, in laundry[2] and dishwashing detergents, cosmetics, food processing[3], skin care ointments[4], contact lens cleaners, and for research purposes in synthetic organic chemistry. In molecular biology using B. subtilis as a model organism, the gene encoding subtilisin (aprE) is often the second gene of choice afteramyE for integrating reporter constructs into, due to its dispensability.
2ND MOLECULE : PROLYL AMINO
Purification and Characterization of a Prolyl Aminopeptidase from Debaryomyces hansenii
ABSTRACT
A prolyl aminopeptidase (PAP) (EC 3.4.11.5) was isolated from the cell extract ofDebaryomyces hansenii CECT12487. The enzyme was purified by selective fractionation with protamine and ammonium sulfate, followed by two chromatography steps, which included gel filtration and anion-exchange chromatography. The PAP was purified 248-fold, with a recovery yield of 1.4%. The enzyme was active in a broad pH range (from 5 to 9.5), with pH and temperature optima at 7.5 and 45°C. The molecular mass was estimated to be around 370 kDa. The presence of inhibitors of serine and aspartic proteases, bestatin, puromycin, reducing agents, chelating agents, and different cations did not have any effect on the enzyme activity. Only iodoacetate, p-chloromercuribenzoic acid, and Hg2+, which are inhibitors of cysteine proteases, markedly reduced the enzyme activity. The Km for proline-7-amido-4-methylcoumarin was 40 μM. The enzyme exclusively hydrolyzed N-terminal-proline-containing substrates. This is the first report on the identification and purification of this type of aminopeptidase in yeast, which may contribute to the scarce knowledge about D. hansenii proteases and their possible roles in meat fermentation.
3RD MOLECULE : REPRESSOR LEX A
DNA damage can be inflicted by the action of antibiotics. Bacteria require topoisomerases such as DNA gyrase or topoisomerase IV for DNA replication. Antibiotics such as ciprofloxacinare able to prevent the action of these molecules by attaching themselves to the gyrase - DNA complex. This is counteracted by the polymerase repair molecules from the SOS response. Unfortunately the action is partly counterproductive because ciprofloxacin is also involved in the synthetic pathway to RecA type molecules which means that the bacteria responds to an antibiotic by starting to produce more repair proteins. These repair proteins can lead to eventual benevolent mutations which can render the bacteria resistant to ciprofloxacin. Mutations are traditionally thought of as happening as a random process and as a liability to the organism. Many strategies exist in a cell to curb the rate of mutations. Mutations on the other hand can also be part of a survival strategy. For the bacteria under attack from an antibiotic, mutations help to develop the right biochemistry needed for defense. Certain polymerases in the SOS pathway are error-prone in their copying of DNA which leads to mutations. While these mutations are often lethal to the cell, they can also lead to mutations which improve the bacteria's survival. In the specific case of topoisomerases, some bacteria have mutated one of their amino acids so that the ciproflaxin can only create a weak bond to the topoisomerase. This is one of the methods that bacteria use to become resistant to antibiotics. Impaired LexA proteolysis has been shown to interfere with ciprofloxacin resistance.[1] This offers potential for combination therapy that combine quinolones with strategies aimed at interfering with the action of LexA either directly, or via RecA.
