A sperm motility inhibitor isolated from porcine seminal plasma is identical to porcine L L-microseminoprotein (MSP). Circular dichroism (CD) and nuclear magnetic resonance (NMR) data showed that the native and recombinant porcine MSPs exhibit very similar structure. The ¢ve disul¢de pairings on porcine MSP were unambiguously assigned based on NMR data and further con¢rmed using structural calculations. Surprisingly, our derived pairings di¡er from those recently reported for ostrich MSP based on matrix-assisted laser desorption ionization-time of £ight (MALDI-TOF) analysis. Furthermore, the secondary structure was determined to comprise one four-stranded and two double-stranded antiparallel L L-sheets. As we know, this is the ¢rst detailed secondary structure reported among several types of MSPs. ß