The protein CDC42 is important for progression of the cell division cycle. This
ID: 65581 • Letter: T
Question
The protein CDC42 is important for progression of the cell division cycle. This link shows the amino acid
sequence of a specific form of CDC42 encoded in human cells:
http://www.ncbi.nlm.nih.gov/nuccore/89903011 Scroll to the bottom and find the nucleotide sequence of
the cDNA encoding this protein, and several dozen lines above it locate the translation (predicted amino
acid sequence) of CDC42. By convention, the base sequence is printed in a 5’ to 3’ direction, and the
amino acid sequence is printed in an N-terminal to C-terminal direction.
Suppose you engineer a chimeric version of Ras, in which its C-terminal 20 amino acids are removed and replaced by the 20 C-terminal amino acids of CDC42. Would this chimeric protein be recognized and lipid-modified by franesyl:protein transferase? Explain your answer briefly for full credit.
Suppose your colleague finds new experimental evidence that CDC42 is a target of a kinase called BTK.
For information on this kinase, see: http://www.uniprot.org/uniprot/Q06187 How many sites of possible
phosphorylation by BTK exist within CDC42? __________
In the primary structure of CDC42, write the two longest regions of consecutive nonpolar amino acid side
chains (not including Cys). ______________________________ __________________________________
What does your answer to the previous question suggest regarding the likelihood that CDC42 harbors a
transmembrane domain?
Explanation / Answer
Protein palmitoylation is a dynamic process that regulates membrane targeting of proteins and protein-protein interactions. We have previously demonstrated a critical role for protein palmitoylation in platelet activation and have identified palmitoylation machinery in platelets. Using a novel proteomic approach, Palmitoyl Protein Identification and Site Characterization, we have begun to characterize the human platelet palmitoylome. Palmitoylated proteins were enriched from membranes isolated from resting platelets using acyl-biotinyl exchange chemistry, followed by identification using liquid chromatography-tandem mass spectrometry. This global analysis identified > 1300 proteins, of which 215 met criteria for significance and represent the platelet palmitoylome. This collection includes 51 known palmitoylated proteins, 61 putative palmitoylated proteins identified in other palmitoylation-specific proteomic studies, and 103 new putative palmitoylated proteins. Of these candidates, we chose to validate the palmitoylation of triggering receptors expressed on myeloid cell (TREM)–like transcript-1 (TLT-1) as its expression is restricted to platelets and megakaryocytes. We determined that TLT-1 is a palmitoylated protein using metabolic labeling with [3H]palmitate and identified the site of TLT-1 palmitoylation as cysteine 196. The discovery of new platelet palmitoyl protein candidates will provide a resource for subsequent investigations to validate the palmitoylation of these proteins and to determine the role palmitoylation plays in their function.
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