Trypsin can be immobilized on silk fibroin fiber (SFF) by introducing several spacer arms, such as ethylene diamine (ED), bovine serum albumin (BSA) and silk sericin (SS). Direct immobilization on silk fiber (SFFGA) has low activity because of the steric hindrance between the trypsin and substrate. The introduction of spacer arms onto SFF-GA can enhance the activity of trypsin by reducing the steric hindrance. When ED is used as a spacer arm, the activity of trypsin has increased but its stability decreased due to the increased hydrophobicity of SFF. BSA and SS, as a spacer arm, have better results in both activity and stability. SFF-BSA shows some decrease in the specific activity due to improper immobilizatin. SFF-SS maintained 90% of its initial activity even after 12 hrs incubation at $50^{circ}C$. In the case of repeated hydrolysis of silk sericin with immobilized trypsin, SFF-GA and SFF-ED lost 50% of their initial activity right after first run, whereas SFF-BSA and SFF-SS maintained 80% of their initial activities even after 5 runs. Higher operational stability is due to increased hydrophilicity of SFF by introducing hydrophilic spacer arms such as BSA and SS. The high content of serine in SS increases the hydrophilicity of SFF resulting the best results among other spacer arms.