Passive vs. active transport Passive transport is simply transport down an electrochemical gradient until equilibrium is reached Active transport results in accumulation of solute beyond equilibrium, the unfavorable thermodynamics is driven by ATP hydrolysis

Passive vs. facilitated diffusion

Four types of transport ATPases F-type – you are familiar with P-type V-type Multi-drug transporter (ABC Transporter)

P-type ATPase Cation transporter that is reversibly phosphorylated as part of the transport cycle Vanadate sensitive Na+, K+, Ca++ Bacteria use to detoxify heavy metals Widely distributed

A P-type ATPase maintains potassium and sodium gradient

This transport mechanism maintains a membrane potential

Ion gradients provide energy for secondary active transport Ion gradients formed by transport of cations can be driving force for cotransport of other solutes

For example,

A composite look at transport

V-type ATPase and Multi-drug transporter V-type –Works as a proton pump –Has key role in acidification of cellular compartments (including endosome) Multi-drug transporter –Export numerous compounds in ATP- dependent manner

Ion channels are distinct from ion transporters Ion channels provide a faster rate of transport Ion channels cannot be saturated Channels are gated, meaning they are open or closed in response to allosteric effectors

Channel structure provides insight into specificity and rate

Features of K + channel Negatively charged amino acids act as sink for cations Pathway narrows (filter) to accommodate interactions specific for potassium Appears to be a paradigm for ion channels (calcium, etc.)

Voltage gated sodium channels Change in membrane potential results in conformational change

Lastly, Ligand-gated channels Acetylcholine receptor binds acetylcholine causing a conformational change opening the ion channel. There are also intracellular ligand-gated channels

Ligand gated ion channels Nicotinic acetylcholine receptor transports sodium, calcium and potassium ions through conformational changes

Membrane proteins such as channels, receptors have significant metabolic roles Hormones and metabolites offer signaling or communication mechanisms within the cell

Qualities of Signal Transduction

Molecular cascades

Feedback inhibition

Integrated networks

Six types of signal transducers

Other two receptor types Receptors with no intrinsic enzyme activity (can interact with enzymes though such as tyrosine kinases) to affect gene expression Adhesion receptor, binds molecules in the extracellular matrix

Looking at receptor-ligand interactions Experimentally must account for non- specific binding (ie. to the membrane, tube, etc.)

Scatchard analysis (like Lineweaver Burke)

Case study: Receptor Enzymes Most commonly – tyrosine kinases

Insulin receptor and a regulatory cascade

Important facets of this process Phosphorylation alters protein structure/function SH2 domain observed in many proteins, a conserved domain that mediates protein- protein interactions G-protein activates kinases Kinases act in a cascade to modulate transcriptional regulators (kinome)

IRS-1 can interact with other cellular components for network integration

G-proteins and signal transduction  -adrenergic Signal pathway

How? Epinephrine binds the serpentine receptor, causing GTP to replace GDP on the G- protein (this particular G-protein is distinct from Ras family) G becomes active with GTP bound, and activates adenyl cyclase, which converts ATP to cAMP Timing mechanism turns off G protein

cAMP activated protein kinases Protein kinase A

Result of epinephrine cascade

Densensitization by phosphorylation

Second messenger cAMP

Other second messengers Diacylglycerol Inositol triphosphate Calcium