Abstract
We recently determined that fusion proteins containing tyrosine-based endocytic signals bind to the mu 2 subunit of AP-2, the complex that drives clathrin coat formation and mediates endocytosis from the plasma membrane. Here we analyze the selectivity of peptide recognition by mu 2 and by AP-2 using combinatorial selection methods and surface plasmon resonance. Both mu 2 and AP-2 are shown to interact with various sequences of the form tyrosine-polar-polar-hydrophobic (Yppø) found on receptors that follow the clathrin pathway. The optimal sequence for interaction with mu 2 and with AP-2 has tyrosine as an anchor and prefers arginine at position Y + 2 and leucine at position Y + 3. In contrast, no preferred sequence is detected surrounding the Yppø signal, indicating that recognition of the Yppø endocytic signal does not require a prefolded structure. We conclude that sorting into the endocytic pathway is governed by a surprisingly simple interaction between the mu 2 chain and a tyrosine-containing tetrapeptide sequence.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Sequence
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Clathrin / chemistry
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Clathrin / genetics*
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Clathrin / physiology*
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / genetics*
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DNA-Binding Proteins / physiology*
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Endocytosis / genetics
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Endocytosis / physiology*
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Humans
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In Vitro Techniques
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Molecular Sequence Data
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Peptide Library
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Protein Conformation
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Protein Sorting Signals / chemistry
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Protein Sorting Signals / genetics*
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Protein Sorting Signals / physiology*
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / physiology
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / physiology
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Signal Transduction / genetics
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Signal Transduction / physiology
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Transcription Factor AP-2
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Transcription Factors / chemistry
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Transcription Factors / genetics*
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Transcription Factors / physiology*
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Tyrosine / chemistry
Substances
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Clathrin
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DNA-Binding Proteins
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Peptide Library
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Protein Sorting Signals
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Recombinant Fusion Proteins
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Transcription Factor AP-2
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Transcription Factors
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Tyrosine