Protein biochem

Proteins

• Complex biopolymers, whose structure is determined by the sequence of amino acids

Alpha Amino Acids

• Contain an asymmetric carbon
• Two stereoisomers exist called enantiomers or optical isomers
• Proteins in all life only contain L-amino acid enantiomers
• The surface of proteins is asymmetric, allowing for highly specific molecular recognition
• Play an important role in the formation of protein “secondary structure”

Methionine and Cysteine

• Both contain sulfur
• Methionine is always the first amino acid in a polypeptide chain
• The free sulfhydryl group of cysteine is highly reactive and is often used in enzyme active sites to attach various chemical groups to proteins
• Cysteine can form disulfide bonds

Primary Protein Structure

• The primary structure of a peptide or protein is the linear sequence of amino acid residues from the N- to the C-terminus.

Secondary Protein Structure

• Dependent on hydrogen bonding involving the peptide bonds
• Alpha Helix
• Beta Sheets
• U Turns
• The secondary structure of a protein is the folding pattern of the polypeptide chain.

Tertiary Structure

• Specifies the special arrangements of the secondary arrangement

Quaternary Protein Structure

• Several subunits held together by non-covalent bonds

Hemoglobin

• One example of a protein with quaternary structure

Sickle Cell Disease

• Abnormal primary structure can affect protein function
• Point mutations leading to misfolding
• Point mutations leading to trapping of protein in the ER
• Point mutations leading to premature stop codons

Insulin

• Proinsulin contains the amino acid sequence of insulin plus the 31-amino acid C (connecting) peptide
• In the Golgi apparatus, disulfide bonds are established, aided by C-peptide
• Sorting and packaging due to special surface proteins
• Insulin is stored in vesicles in zinc-bound crystals
• Proteases that cleave proinsulin are packaged within the secretory vesicle

Amyloidosis

• Accumulation of amyloid (proteins that have folded abnormally and aggregated together)
• Folded into beta sheets, they form extracellular deposits

Prions

• Proteinaceous infectious particles
• PrPC (Prion Protein) is a normal neuronal protein involved in cell adhesion, ion channel activity, and neuronal excitability
• In abnormal form, alpha helix changes to beta sheet
• Abnormal PrPC can convert normal PrPC into abnormal form

Histopathology

• Creutzfeldt-Jakob Disease (CJD)
• Gerstmann-Straussler-Scheinker Syndrome
• Kuru
• Scrapie in sheep
• Bovine Spongiform Encephalopathy (BSE) in cows
• Transmissible Mink Encephalopathy in mink
• Feline Spongiform Encephalopathy in cats

Protein Detection

• SDS Page gels – 1D and 2D
• Staining techniques
• Immunoblotting
• Mass spectrometry
• Crystallography
• Edman sequencing

Ubiquitination

• Ubiquitin is an 8.5kDa protein
• Marks proteins for degradation via the proteasome
• Binds to lysine, cysteine, threonine residues along the length of the target molecule or to the N-terminus of the protein
• Regulates major cellular processes such as cell division, immune responses, and embryonic development

Protein Breakdown

• Important way for cells to regulate expression of proteins
• The proteasome

Functions of Proteins

• Structural proteins
• Catalytic proteins – Enzymes
• Signaling Proteins
• Proteins involved in cell adhesion and recognition
• Membrane transport proteins

Functions of Proteins: Examples

• Extracellular Matrix Proteins (ECM) like collagen, Elastin, lamin and fibronectin
• Muscle proteins: actin and myosin
• Cytoskeletal proteins
• Proteins that participate in immunity (antibodies, complement system…)
• Transport proteins including Albumin (50% of plasma protein)
• Pro-and anti-coagulant proteins
• DNA binding proteins

Summary

• Proteins are organized in four different levels (primary to quaternary structure)
• There are several different detection levels for proteins
• Proteins can be post-translationally modified
• Proteins fulfill a wide array of functions