Introduction+to+Proteins+III+Part+1


 * 22 August 2006**
 * Introduction to Proteins III Part 1**
 * Dr. Kandace Williams, Ph.D.**

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=**3-D Structures**=


 * Primary structure is the amino acid sequence
 * [|Secondary structures] are [|α-helices] and [|β-pleated sheets]
 * Tertiary structure is one complete protein chain
 * Quaternary structure are for separate chains assembled into an oligometric protein

**Tertiary Structure**

 * Tertiary structure of proteins are non-symmetrical
 * Spatial arrangement of amino acid residues within a polypeptide chain that are far apart in sequence, including pattern of disulfide bonds
 * Form by **thermodynamic stability** and **contribution of each portion to overall polypeptide chain-folding mechanisms**

**Myoglobin**

 * One [|myoglobin] protein contains 153 amino acids
 * MW is 16.83 kDaltons
 * ~70% composed of 8 α-helixes; remainder composed of turns and loops, forming a compact globular shape (much larger when denatured)
 * Overall shape is devoid of symmetry
 * Interior contains non-polar residues
 * except for two histidines to bind iron and oxygen
 * Exterior contains both polar and non-polar residues

**Hydrophobicity and Hydrophilicity**

 * Water soluble proteins are amphipathic – meaning both polar and non-polar
 * Generally fold in compact structures with non-polar cores
 * Hydrophilic –R groups (polar) --> Outside
 * Hydrophobic –R groups (non-polar) --> Inside
 * Backbone made hydrophobic by hydrogen bonding within α-helix and β-pleated sheets
 * Polar –R groups made hydrophobic by van der Waals interactions.
 * Exception: Porin within outer membrane of bacteria is hydrophobic outside (interact with lipid membrane) and hydrophilic inside (allows polar molecules through)

**Protein Domains**

 * Two or more compact globular protein regions connected by flexible segment of polypeptide chain
 * Protein Domains are within the same protein; do **not** confuse with quaternary structure which involves more than one protein

=**Quaternary Structure**=


 * Spatial arrangement of proteins containing more than one polypeptide chain
 * Each protein in a quaternary structure is called a subunit
 * Dimmers contain two identical subunits
 * Tetramer consists of four subunits

=**Protein Folding**=


 * Denature protein using:
 * [|β-mercaptoethanol] reduces covalent disulfide bonds to sulfhydryls (-S-S- to –SH + HS-)
 * Strong reducing agent
 * Urea and guanidinum chloride can disrupt hydrogen bonds and hydrophobic interactions
 * Denaturing protein using β-mercaptoethanol, urea and guanidinum chloride destroys enzymatic activity
 * Protein activity can be recovered when β-mercaptoethanol, urea and guanidinum chloride were removed and oxygen added suggesting correct refolding of the protein with correct disulfide bonds reforming
 * if only β-mercaptoethanol removed but keep urea and guanidinum chloride, protein can reform but disulfide bonds form randomly and is enzymatically inactive
 * Randomly reformed disulfide bonds inhibit proper refolding
 * Proteins have inherent ability to form its correct tertiary structure
 * “If you leave it alone, it’ll figure out what its supposed to do”
 * Primary structure specifies conformation of the final tertiary protein
 * Protein folding in tertiary structure is not a random processes
 * Levinthal’s paradox – calculated versus actual time for a protein to fold correctly
 * Predict that it would take 1.6 x 10 e27 years to fold correctly by random search
 * Actually, E. coli produces 100 active proteins in 5 seconds
 * Cooperative transition in protein folding – “all or none”
 * Partially denatured solution has 50% folded and 50% unfolded proteins
 * Sharp transition between folding native protein and denatured unfolding protein (and vice versa)
 * Because unfolding proteins form aggregates and takes up space, it is important for proteins to have an “all or none” approach to protein folding to avoid forming aggregates
 * Chaperones help proteins to correctly fold and avoid aggregates
 * Cumulative selection aids protein folding
 * Retention of partially correct intermediates by differences in free energy and leads to increasing stability as the entire correct conformation is achieved
 * That is, as soon as the first fold is correct, it helps the next fold being correct

=**Prediction of 3-D protein structures**=


 * ab initio predictions
 * Predict secondary and tertiary structures based by amino acid sequence without reference to known protein structures
 * Computers used to determine differences in free energy of protein structures
 * Utility limited by sheer number of possible configurations
 * Knowledge-based methods
 * Amino acid sequence of unknown protein compared for sequence compatibility with proteins of known conformation by computer analysis
 * If significant match is found, it is used as an initial model
 * Predictions of secondary structures with 6 or fewer residues are 60-70% accurate
 * Conformational preferences of amino acids are **not** absolute
 * Tertiary interactions and microenvironment also play roles not fully understood

=**Human Disease**=
 * Several neurogenerative diseases are disease of protein misfolding.
 * [|Prions] – self-propagating
 * Infectious protein replicates by transferring protein misfolding in the absence of nucleic acid from α-helices to form insoluble aggregates of β-pleated sheets
 * Prions are responsible for transmissible spongiform encephalopathies (TSE)
 * Fatal neurodegenerative disorders affecting humans and other mammals
 * Human = [|Creutzfeldt-Jakob disease] (CJD) diagnosed in ~1/million/year
 * Bovine = [|Bovine spongiform encephalopathy] (BSE)
 * Human CJD is 85% sporadic, 10-15% is inherited (fCJD), and <5% are infectious (iCJD)
 * Iatrogenic (physician-caused) iCJD transmitted by medical or surgical, human material injected, etc.
 * Variant vCJD – Can BSE be transferred to CJD

=**Objectives**=