This study reports on new proteolytically stable, pharmacologically active endomorphin analogues, incorporating Dmt(1), Achc(2), pFPhe(4), or βMePhe(4) unnatural amino acids. Consistent with earlier results, it was found that the analogues carrying Dmt(1) and Achc(2) residues displayed the highest μ-opioid receptor affinities, depending upon the configuration of the incorporated Achc(2). Combination of such derivatives with pFPhe(4) or βMePhe(4) yielded further compounds with variable binding potencies. Combined application of Dmt(1), cis-(1S,2R)Achc(2), and pFPhe(4) (compound 16) resulted in the most potent analogue. Ligand stimulated [(35)S]GTPγS binding assays indicated that the analogues retained their agonist activities and opioid receptor specificities. NMR and molecular modeling studies of the analogues containing βMePhe(4) or pFPhe(4) confirmed the predominance of bent structures, however, it is apparent that bent structures are energetically more favored than random/extended structures for all studied compounds.