In this study novel miniprotein inhibitors against the human type II transmembrane serine protease matriptase (MT-SP1) should be identified. Therefor the two different methods of protein engineering, rational protein design and directed evolution have been applied to the miniprotein scaffold. For the directed evolution approach the technique of yeast surface display has been used. First the suitability of the natural occurring miniprotein variants of MCoTI as inhibitors against matripase has been tested. The miniprotein variants have been isolated from the seeds of Momordica cochinchinensis. Next the inhibition constants of those variants against the catalytic domain of matriptase, which has been produced and purified from E.coli, were determined. It turned out that MCoTI-II with a Kiapp of 235 nM is the best variant and therefore it has been used as a scaffold for the identification of novel miniprotein inhibitors in the following parts of this study. In the next step different positions for an amino acid exchange in the miniprotein have been identified with the rational protein design method. The amino acids exchanges were introduced into the miniprotein via solid phase peptide synthesis to obtain variants with higher affinity against matriptase. Determination of the inhibition constants showed that these experiments did not yield miniprotein variants with a higher affinity against matriptase. In the next step the directed evolution method has been applied to MCoTI-II. Therefor two combinatorial libraries, with diversities of 1E+7 and 2E+7, have generated in S.cerevisiae. These two libraries have been sorted against decreasing concentrations of matriptase using the technique of yeast surface display and fluorescence assisted cell sorting (FACS). A subset of miniprotein variants from these selections has been produced via solid phase peptide synthesis and their inhibition constants have been determined. From this method of protein engineering two novel miniprotein inhibitors against the human type II transmembrane serine protease matriptase with high affinities (Kiapp 3,7 nM and 8,7 nM) could be identified.