MicroRNAs (miRNAs) are a class of 19-25 nt long non-coding RNAs that regulate gene expression post-transcriptionally by binding with partially complementary sequences in the T-untranslated region (3'-UTR) and inhibiting mRNA translation or by affecting mRNA stability. We have characterized the structures and the equilibrium between hairpin and homo-duplex form of the mature strand of hsa-mir-520h by various concentration and temperature dependent 1D, 2D NMR experiments and those structures correspond well with Mfold-folded and UNAFold-aligned secondary structures. A detailed folding and alignment analysis in physiological conditions of all mature miRNA strands from the complete database of known miRNAs (miRBase) was performed. The statistical analysis of the resulting folding and alignment data showed for the first time the potential of a large number of mature miRNAs to form significant hairpin and/or homo-duplex structures in solution. The self-complementarity of mature miRNAs can provide a mechanistic tuning and a regulatory sophistication to the process of miRNA mediated gene regulation. (C) 2010 Elsevier Inc. All rights reserved.