What Can Studies Of Serine Proteases Tell Us About The Relationship Between Protein Structure, Function And Evolution?

5. Explain why proteases are secreted in an inactive state and describe the means by which proteases are activated in the stomach and small intestine. – The enzymes protease, amylase and lipase, are potent enzymes that are capable of digesting the pancreatic cells- a process called autodigestion. To protect themselves from autodigestion in case the digestive enzymes accidentally get turned on, pancreatic cells produce enzymes in inactive form called zymogens, which are stored in membrane-bound sacks called zymogen granules. Enteropeptidase (also called enterokinase) is an enzyme produced by cells of the duodenum

Proteins with his-tags are purified using IMAC, immobilized metal affinity chromatography. The protein product interacts with a metal that is reversibly bound to an immobilized chelating group. The immobolized chelating group acts as a Lewis base (electron-pair donor) to which the Lewis acid (electron-pair acceptor) metal ion is coordinated. The support to which the metal ion binds is called a ligand. When an electron donor group is replaced by another, the action is referred to as ligand exchange. The donor atoms involved in this exchange are the electronegative nitrogen, sulfur, or oxygen. These atoms scavenge for sources of electrons. The structure formed when the metal ions are added to form the chelate result in free coordination

is resistant to protease and as a result, these prions replicate and are not broken

Reports indicate that the extracellular serine protease inhibitor, C1INH, can effectively bind HABP2 in the blood (27). We therefore examined whether C1INH can colocalize with HABP2 in

Basic secretions from the pancreas then enter the small intestine to neutralize the pH-decreasing effect of the acidic chyme. Thus these basic and acidic fluid secretions in the duodenum, cause the pH in the duodenum to fluctuate from an acidic pH of 2 to a basic pH of 7.5 (Woodtli & Werner, 1995). We wanted to investigate the question of how these fluctuations of pH in the duodenum affect the activity of the enzyme Trypsin. Enzymes play a very important physiological role in the human body because they speed up the rates of reactions vital to our survival. Enzymes bind to specific substrates and configure their positions, geometry or interactions relative to one another in such a way, that the activation energy required for a reaction to occur is lowered, and thus the reaction rate is increased. The activity of an enzyme depends on many environmental factors such as pH and temperature, because changes in temperature and pH affect the intermolecular interactions between amino acids in the backbone, holding the tertiary structure of the enzyme together. Thus each enzyme has an optimal pH range, below which, and above which its activity declines. Trypsin is an important enzyme that catalyzes the breakdown of specific peptide bonds in the duodenum of the small intestine so that we can extract nutrients and energy from the proteins we eat (Nelson & Cox, 2013). Trypsin is a

Proteins are complex macromolecules which are essential for life of all organisms. They are manufactured through the processes of transcription and translation, which take place inside the cells. More specifically, they are synthesised by ribosomes (Shakhnovich, 2007). Figure 1.1 shows the overall processes that can occur in making a fully functionally active protein. (Ghelis, 2012). Functional properties of certain proteins include, but is not limited to: structural composition of the cytoskeleton in cells, catalysing biochemical reactions and hormones such as insulin to regulate blood sugar concentration (O’Connor & Adams, 2014). Proteins must therefore have a specific 3-dimentional shape to allow them to carry out these functions. If their

Chymotrypsin is a digestive enzyme component of pancreatic juice acting in the duodenum where it performs proteolysis, the breakdown of proteins and polypeptides. Chymotrypsin preferentially cleaves peptide amide bonds where the carboxyl side of the amide bond (the P1 position) is a largehydrophobic amino acid (tyrosine, tryptophan, and phenylalanine). These amino acids contain an aromatic ring in their sidechain that fits into a 'hydrophobic pocket' (the S1 position) of the enzyme. It is activated in the presence of trypsin. The hydrophobic and shape complementarity between the peptide substrate P1 sidechain and the enzyme S1 binding cavity accounts for the substrate specificity of this enzyme. Chymotrypsin also hydrolyzes other amide bonds in peptides at slower rates, particularly those containing leucine and methionine at the P1 position.

In the present study, 2 types of mutations were observed with different frequencies. Amino acid frequencies at each position in the NS3 protease sequence were determined with the VESPA software program. 24 Genotype 4 -specific amino acid signatures were present in almost all of our

The proteins produced are mainly to be used within the cell and the enzyme which are products by proteins help in speeding up certain biological processes.

Proteins are the most abundant organic matter in living organisms. They have several functions such as; Structure, Movement, Transport, Buffering, Metabolic activity, Communication and Defence/Protection. (Creighton, 1989). Proteins are assembled using different structures known as Primary structures to Quaternary structures. The Primary structure of the protein is the sequence or order of amino acids in the polypeptide chain. Once several amino acids have joined together in a chain, the chain tends to make certain shapes or patterns. Peptide bonds contain polar hydrogen atoms (with a small positive charge) and polar oxygen atoms (with a small negative charge). This allows hydrogen bonds to form between peptide bonds in different parts of the chain. Because of this, the polypeptide chain can take on different shapes or patterns in different parts of the chain, and these patterns are called the Secondary Structure of the protein. The two main types of secondary structure in proteins are the alpha helix and the beta pleated sheet. The secondary structure of the protein depends on the amino acids it contains, and the order in which they join together. In other words, it depends on the primary structure. The Tertiary Structure of a protein is the overall three-dimensional shape. It has a specific shape and is held together by weak bonds between the side chains (R groups) of the different

Proteins are arguably the most structurally and functionally complex structures known to man. Through evolution spanning billions of years, these macromolecules have been perfected to perform many functions necessary for life. Proteins contain four distinct levels of structure and are composed of simple organic compounds known as amino acids. (Alberts, Bruce et al 2000) Amino acids are joined together by peptide bonds made during protein biosynthesis, and the linked amino acid chain is known as a polypeptide chain. DNA determines the amino acid sequence and the sequence is what determines the primary level of protein structure. This level of structure determines the protein’s shape, which affects the assembly and function of

Studying the molecular basis of Gag-protease coevolution, Ozen et al. found that coevolved Gag-protease complexes were also able to restore protease active site dynamics by increasing Gag-protease interactions. Under selective pressures of PIs, Ozen et al. found that

The active site function was the same for both. Based on Fig. 1, the active site for Caspase-1 is His237 and Cys285 and for Caspase-4 isoform alpha the active site is His209 and Cys257. For Caspase-1, the active site residues His209 is important because as a base it demonstrates nucleophilic behavior when attracting a proton from Cys285. Thus, His209 acts as a proton shuttle. For the active site residue, Cys285, it acts as a nucleophile; this cysteine is important for cleaving the peptide bond in proteins that contain aspartic acid. To further characterize the enzyme Caspase-1, the properties of the protein were compared to the homologous protein Caspase-4 isoform alpha (Table 1).

The fluorescence of RepA70-YFP increased as more RepA70-CFP was produced (Fig. 3C), and this indicated that ClpAP protease could be overloaded and a proteolytic-queue forms similar to what was observed with the LAA tagged proteins targeted to ClpXP8. We also tested two other tags, MarA and MarAn20 (20 amino acids from the N-terminal of MarA), which target proteins to be degraded by the Lon protease. The Lon protease was weakly overloaded by MarA tagged proteins but was overloaded more by MarAn20 tagged proteins (Fig. 3C-D). This made us wonder if Lon could be overloaded when both MarA and MarAn20 were co-produced. Indeed, this was the case (Fig. 3E).

The analysis of a complete set of proteins (proteome) of a given cell or organism can be defined as proteomics (Phillips & Bogyo, 2005). The primary aim of proteomic analysis is to separate, identify and characterize proteins and understand their interactions with other proteins. There are four branches of proteomics which are sequence and structural proteomics, expression proteomics, interaction proteomics and functional proteomics.