Molecular Mechanisms of Ion Channel Function 
Gary Yellen, Ph.D. — Assistant Investigator 
Dr. Yellen is also Assistant Professor of Neuroscience and Biophysics at the Johns Hopkins University 
School of Medicine. He received his undergraduate degree in biochemical sciences from Harvard College 
and his Ph.D. degree in physiology from Yale University, where he studied with Charles Stevens. Dr. Yellen 
did his postdoctoral research on ion channel physiology as a Life Sciences Research Foundation postdoc- 
toral fellow at Brandeis University, where he worked with Christopher Miller. 
ALL electrical signaling in the nervous system 
is controlled by ion channels, a class of 
membrane proteins that form pores through the 
membrane. Charged ions such as sodium, potas- 
sium, and calcium pass through ion channels and 
carry an electrical current. The channels them- 
selves are regulated, so that the pores are only 
open when the proper chemical or electrical sig- 
nal is present, and only certain ions can pass 
through a particular kind of channel. By under- 
standing how^ channels open and close and how 
they are regulated, we define the repertoire of 
molecular changes used by neurons when they 
signal, sense, and learn. 
Ion channels, like other membrane proteins, 
have resisted standard biochemical and structural 
analysis. Their structure has only recently begun 
to be elucidated by a combination of protein 
chemistry and molecular biology. On the other 
hand, we have detailed knowledge of the func- 
tioning of ion channels. Because each ion chan- 
nel catalyzes the transport of millions of ions per 
second, we can measure electrically the current 
carried by just a single-channel protein molecule. 
This technique of single-channel recording has 
allowed us to make a detailed model for the con- 
formational changes between open and closed 
states induced by chemical ligands and changes 
in voltage, but we still have no knowledge of the 
protein structures that underlie these conforma- 
tional changes. 
My laboratory is using two complementary ap- 
proaches to relate ion channel function to struc- 
ture. Site-directed mutagenesis allows us to mod- 
ify any amino acid in a protein for which we have 
the cloned genetic material. Rather than modify- 
ing the protein directly, we change the DNA se- 
quence and then inject the modified messenger 
RNA into immature frog eggs (oocytes), which 
manufacture the modified protein. This method 
allows us to test specific theories about which 
parts of the channel protein are important for spe- 
cific functional features. 
We have applied this first strategy to voltage- 
activated potassium channels, which participate 
in electrical signaling. By systematic mutagene- 
sis, we have identified the region of the potas- 
sium channel protein that lines the pore through 
which ions cross the membrane. We found spe- 
cific amino acid residues in the protein sequence 
that control the sensitivity of the channel to tet- 
raethylammonium, an organic ion that can block 
current through the channel. Natural variation of 
one of these amino acids explains the differences 
in drug sensitivity between potassium channels 
in different organs or species. Amino acids in this 
region of the protein can also affect the rate at 
which ions are conducted through the pore. 
These discoveries put us in a position to discover 
the basic mechanisms of ion selectivity and chan- 
nel gating at the level of individual amino acids. 
We have also used this strategy to study acetyl- 
choline-activated cation channels, which convert 
neurochemical signals into electrical signals at 
synapses. We have changed amino acids in the 
region of the protein that binds acetylcholine and 
identified specific residues that play a critical 
role in binding and signal transduction by acetyl- 
choline. These studies are teaching us more 
about the molecular basis of drug recognition 
and signal transduction in this protein. 
A second approach for finding parts of the pro- 
tein that are important for specific functions in- 
volves mutant selection. We are developing a sys- 
tem in which we can randomly mutagenize the 
gene for a channel and then select the interesting 
mutants. This strategy is almost the reverse of 
site-directed mutagenesis. Instead of choosing to 
change a particular part of the protein and look- 
ing for the resultant changes in function, we se- 
lect for a change in function and then look at the 
underlying structural changes. We have ex- 
pressed the genes for the nicotinic acetylcholine 
receptor in tissue-cultured mammalian cell lines 
in preparation for this mutant selection strategy. 
We have also found a method for selecting certain 
channel mutants by using a fluorescently tagged 
toxin molecule that binds differently to these 
mutant channels. 
491 
