Characterization　of　the　protein-peptide　interactions　are　a　critical　for　understanding　the　functions　and　signal　pathways　of　proteins.　Herein,　a　new　finding　of　universal　terminal　protection　that　protein　bind　specifically　with　peptide　and　provide　a　protective　coating　to　prevent　peptide　hydrolysis　in　the　presence　of　peptidase.　On　the　basis　of　this　mechanism,　we　first　reported　a　novel　label-free　fluorescence　biosensor　strategy　that　utilizes　the　protection　of　specific　terminal　protein　on　peptide-templated　gold　nanocluster　(AuNCs)　beacon　for　the　detection　of　proteins.　The　fluorescence　quenching　of　peptide-templated　AuNCs　can　be　effectively　inhibited　with　increasing　concentration　of　the　specific　protein,　exhibiting　a　satisfactory　sensitivity　and　selectivity　toward　protein　with　the　detection　limit　of　MDM2　and　gp120　are　0.0019　U/mL　and　0.0012　U/mL,　respectively.　The　developed　label-free　fluorescence　biosensor　strategy　provides　new　ideas　to　detect　and　screen　protein　for　analyzing　protein-peptide　interaction　in　biomedical　applications.　©　2021　Elsevier　B.V.