The crystal structure, thermal vibrations, and electron density of leu-enkephalin trihydrate, C28H37N5O7.3H(2)O, have been analyzed using single-crystal X-ray diffraction data measured at 100 K with Mo K alpha radiation to a resolution corresponding to [(sin theta)/lambda](max) = 1.15 Angstrom(-1). A multipolar atom density model was fitted against the 5444 unique data with I > 2 sigma(I) [R(F) = 0.038, R(w)(F) = 0.023, GOF = 0.73] in order to calculate the phases for the crystal structure factors and map the valence electron distribution. Ab initio SCF calculations have been carried out on molecular fragments with extended basis sets including polarization functions. The experimental densities along the equivalent bonds of the four peptide groups are equal to within 0.05 e Angstrom(-3) and are in close agreement with those obtained for smaller peptides. Fragment theoretical distributions derived either with or without the molecules of crystallization water are used to figure out the influence of hydrogen bonds on the deformation density. Experimental and theoretical maps derived in the plane containing the intramolecular H bonds characteristic of the beta turns evidence the character of weak N-H...pi bonds of those interactions. Leu-enkephalin trihydrate crystallizes in the orthorhombic space group P2(1)2(1)2(1) With M(r) = 609.68, D-x= 1.34 g cm(-3), Z = 4, and cell dimensions (100 K) a 10.851(4) Angstrom, b = 13.095(3) Angstrom, c 21.192(4) Angstrom, and V = 3011(3) Angstrom(3).