G protein-coupled receptor kinase 2 (GRK2) plays a central role in the cellular transduction network. In particular, during chronic heart failure GRK2 is upregulated and believed to contribute to disease progression. Thereby, its inhibition offers a potential therapeutic solution to several pathological conditions. In the present study, we performed a SAR study and a NMR conformational analysis of peptides derived from HJ loop of GRK2 and able to selectively inhibit GRK2. From Ala-scan and d-Ala point replacement, we found that Arg residues don't affect the inhibitory properties, while a d-amino acid at position 5 is key to the activity. Conformational analysis identified two β-turns that involve N-terminal residues, followed by a short extended region. These information can help the design of peptides and peptido-mimetics with enhanced GRK2 inhibition properties. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 121–128, 2014.