LOCALIZATION OF THE INSULIN-LIKE GROWTH-FACTOR-II BINDING-SITE TO AMINO-ACIDS 1508-1566 IN REPEAT 11 OF THE MANNOSE 6-PHOSPHATE INSULIN-LIKE GROWTH-FACTOR-II RECEPTOR/

The mannose 6-phosphate/insulin-like growth factor II receptor (M6P/IG F-II receptor) binds insulin-like growth factor II (IGF-II) with high affinity. To localize the IGF-II binding site within the 15 repeating units that form the extracytoplasmic domain of the receptor, purified human M6P/IGF-II receptor was digested with thermolysin, and the fragm ents were analyzed for their ability to bind I-125-IGF-II in a cross-l inking assay. Two IGF-II-binding receptor fragments of 23 and 37 kDa w ere purified. Sequence analysis revealed that the fragments consist of disulfide connected peptides comprising amino acids 1331-1566 and 133 1-1697 of the receptor repeats 9-12. In a second approach we expressed truncated forms of the M6P/IGF-II receptor fused to the C terminus of the extracytoplasmic domain of the 46-kDa mannose 6-phosphate recepto r. Fusion proteins containing M6P/IGF-II receptor repeats 10-15, 10-11 , or 11-15 bound IGF-II, whereas a fusion protein containing the singl e repeat 10 failed to bind. This result indicates that repeat 11 (amin o acids 1508-1650) is sufficient for binding of IGF-II. Residues 1508- 1566, which are shared by the 23 kDa IGF-II-binding fragment and repea t 11, are proposed to form the IGF-II binding site of the M6P/IGF-II r eceptor.