Problem Set #10: Membrane Proteins
1. Using the hydropathy plotting program at http://fasta.bioch.virginia.edu/o_fasta/grease.htm, analyze the following membrane proteins:
Protein A: sgfefhgyarsgvimndsgastksgayitpagetggaigrlgnqadtyvemnlehkqtldngattrfkvmvadgqtsyndwtastsdlnvrqafvelgnlptfagpfkgstlwagkrfdrdnfdihwidsdvvflagtgggiydvkwndglrsnfslygrnfgdiddssnsvqnyiltmnhfagplqmmvsglrakdnderkdsngnlakgdaantgvhallglhndsfyglrdgssktallyghglgaevkgigsdgalrpgadtwriasygttplsenwsvapamlaqrskdryadgdsyqwatfnlrliqainqnfalayegsyqymdlkpegyndrqavngsfykltfaptfkvgsigdffsrpeirfytswmdwskklnnyasddalgsdgfnsggews
Protein B: LTTWLWIGAAGMLLGTLLFIYMGRGLEDPEARKIYAATILIPGIAIVAYLGMALGIGVTTVEMPAEHIYWARYLDWLLTTPLLLLFLGLLAGADRRTIFTLVVADALMIVTGLAAALTTSSGLLRWVLFGISTAFFLVLLYYLYVNFPRAAKDAPGTASLFNTLRNLTVVLWLAYPVVWLLGPEGIGLLTVGIETLLYVVLDVVAKVGFGFILLRSRAVLERTVASALAAGS
Please sketch out the hydropathy plot and use it to predict a protein topology (relative to the membrane) for each of the proteins. Now, use Entrez to identify the identity of these membrane proteins, and compare your predictions to the actual structures. Was the hydropathy analysis useful for predicting the topology of these proteins? Why or why not?
2. In class we discussed the potassium channel and the basis of its selectivity for the potassium cation over other cations. For reference, that channel is discussed in your text and in the papers referenced in the class handouts, and can be seen in the structure file 1BL8.pdb.
The structure of the chloride channels from two bacteria have also been solved, revealing the basis of anion selectivity. Look at the original paper (Dutzler et al. Nature, v. 415 p 287) and structure files (1KPK. pdb, 1KPL.pdb). Roughly sketch out the protein structure, indicating its orientation in the membrane, the direction of ion flow, etc. Where is the selectivity filter, and how does it allow Cl- ions, but not other ions, to pass through? Does this selectivity filter share any features with the potassium ion channel?
In the pores through both the potassium and chloride ion channels, do ions ever directly bind to lysines, arginine, glutamates, or aspartates? Why?
3. Why are the structures of membrane proteins difficult to determine? How then have some membrane proteins been crystallized? Please be specific, and reference specific papers. (to start, try your text, the PDB, and http://blanco.biomol.uci.edu/Membrane_Proteins_xtal.html)
4. If you identified a new putative membrane protein but could not crystallize it, how might you go about (a) predicting and (b) characterizing its structure?