MCAT Biochemistry Review
Chapter 3: Nonenzymatic Protein Function and Protein Analysis
Biosignaling is a process in which cells receive and act on signals. Proteins participate in biosignaling in different capacities, including acting as extracellular ligands, transporters for facilitated diffusion, receptor proteins, and second messengers. The proteins involved in biosignaling can have functions in substrate binding or enzymatic activity.
Ion channels are proteins that create specific pathways for charged molecules. They are classified into three main groups, which have different mechanisms of opening, but all permit facilitated diffusion of charged particles. Facilitated diffusion, a type of passive transport, is the diffusion of molecules down a concentration gradient through a pore in the membrane created by this transmembrane protein. It is used for molecules that are impermeable to the membrane (large, polar, or charged). Facilitated diffusion allows integral membrane proteins to serve as channels for these substrates to avoid the hydrophobic fatty acid tails of the phospholipid bilayer. The three main types of ion channels are ungated, voltage-gated, and ligand-gated.
Hundreds of ion channels have been identified that function in cell signaling and cell excitability. In addition, ion channels are drug targets in the treatment of everything from heart disease (calcium channel blockers) to seizures (sodium channel blockers) to irritable bowel syndrome (acetylcholine receptor/cation channel blockers).
As their name suggests, ungated channels have no gates and are therefore unregulated. For example, all cells possess ungated potassium channels. This means there will be a net efflux of potassium ions through these channels unless potassium is at equilibrium.
In voltage-gated channels, the gate is regulated by the membrane potential change near the channel. For example, many excitable cells such as neurons possess voltage-gated sodium channels. The channels are closed under resting conditions, but membrane depolarization causes a protein conformation change that allows them to quickly open and then quickly close as the voltage increases. Voltage-gated nonspecific sodium–potassium channels are found in cells of the sinoatrial node of the heart. Here, they serve as the pacemaker current; as the voltage drops, these channels open to bring the cell back to threshold and fire another action potential, as shown in Figure 3.4.
Figure 3.4. Action Potential of the Sinoatrial Node
For ligand-gated channels, the binding of a specific substance or ligand to the channel causes it to open or close. For example, neurotransmitters act at ligand-gated channels at the postsynaptic membrane. The inhibitory neurotransmitter γ-aminobutyric acid (GABA) binds to a chloride channel and opens it.
The activity at the neuromuscular junction and most chemical synapses relies on ligand-gated ion channels. The nervous system especially makes use of this type of gating, as well as voltage-gated ion channels, as discussed in Chapter 4 of MCAT Biology Review.
The Km and vmax parameters that apply to enzymes are also applicable to transporters such as ion channels in membranes. The kinetics of transport can be derived from the Michaelis–Menten and Lineweaver–Burk equations, where Km refers to the solute concentration at which the transporter is functioning at half of its maximum activity.
Membrane receptors may also display catalytic activity in response to ligand binding. These enzyme-linked receptors have three primary protein domains: a membrane-spanning domain, a ligand-binding domain, and a catalytic domain. The membrane-spanning domain anchors the receptor in the cell membrane. The ligand-binding domain is stimulated by the appropriate ligand and induces a conformational change that activates the catalytic domain. This often results in the initiation of a second messenger cascade. Receptor tyrosine kinases (RTKs) are classic examples. RTKs are composed of a monomer that dimerizes upon ligand binding. The dimer is the active form that phosphorylates additional cellular enzymes, including the receptor itself (autophosphorylation). Other classes of enzyme-linked receptors include serine/threonine-specific protein kinases and receptor tyrosine phosphatases.
Biosignaling can take advantage of either existing gradients (ion channels) or second messenger cascades (enzyme-linked receptors and G protein-coupled receptors).
G PROTEIN-COUPLED RECEPTORS
G protein-coupled receptors (GPCRs) are a large family of integral membrane proteins involved in signal transduction. They are characterized by their seven membrane-spanning α-helices. The receptors differ in specificity of the ligand- binding area found on the extracellular surface of the cell. In order for GPCRs to transmit signals to an effector in the cell, they utilize a heterotrimeric G protein. G proteins are named for their intracellular link to guanine nucleotides (GDP and GTP). The binding of a ligand increases the affinity of the receptor for the G protein. The binding of the G protein represents a switch to the active state and affects the intracellular signaling pathway. There are several different G proteins that can result in either stimulation or inhibition of the signaling pathway. There are three main types of G proteins:
· Gs stimulates adenylate cyclase, which increases levels of cAMP in the cell
· Gi inhibits adenylate cyclase, which decreases levels of cAMP in the cell
· Gq activates phospholipase C, which cleaves a phospholipid from the membrane to form PIP2. PIP2 is then cleaved into DAG and IP3; IP3 can open calcium channels in the endoplasmic reticulum, increasing calcium levels in the cell
Figure 3.5. Trimeric G Protein Cycle (Gs or Gi)
Functions of heterotrimeric G proteins:
· Gs stimulates.
· Gi inhibits.
· “Mind your p's and q's”: Gq activates phospholipase C.
The three subunits that comprise the G protein are α, β, and γ. In its inactive form, the α subunit binds GDP and is in a complex with the β and γ subunits. When a ligand binds to the GPCR, the receptor becomes activated and, in turn, engages the corresponding G protein, as shown in Step 1 of Figure 3.5. Once GDP is replaced with GTP, the α subunit is able to dissociate from the β and γ subunits (Step 2). The activated α subunit alters the activity of adenylate cyclase. If the α subunit is αs, then the enzyme is activated; if the α subunit is αi, then the enzyme is inhibited. Once GTP on the activated α subunit is dephosphorylated to GDP (Step 3), the α subunit will rebind to the β and γ subunits (Step 4), rendering the G protein inactive.
MCAT Concept Check 3.2:
Before you move on, assess your understanding of the material with these questions.
1. Contrast enzyme-linked receptors with G protein-coupled receptors:
2. What type of ion channel is active at all times?
3. How do transport kinetics differ from enzyme kinetics?