Michael D. Lane

     E-mail: lane0280@umn.edu

     Year Entered: 2009

     Degrees Received:
     University of Florida
     Microbiology and Cell Science major
     B.S., 2006

     Honors and Awards:

     -American Heart Association Research Fellowship, 2012-2017

     Thesis Advisor: Burckhard Seelig, PhD

     Thesis Research: Directed evolution of streptopain to inactivate
     superantigens

     Streptococcus pyogenes is a pervasive bacterium capable of causing
     a wide range of human disease, from the simple “strep throat” to the terrifying “flesh eating disease” (necrotizing fasciitis).  When introduced to the bloodstream, S. pyogenes can release extremely potent toxins, named “superantigens,” causing streptococcal toxic shock syndrome (STSS) and death within hours. Proteolytic neutralization of superantigens would prevent these deadly toxicities, but superantigens have shown natural resistance to proteolysis. Recently, streptopain (a cysteine protease produced by S. pyogenes) was shown to degrade SmeZ (the most potent superantigen) yet not SpeA (a superantigen linked to STSS and necrotizing fasciitis) despite over 50% sequence identity and a very similar fold. We will evolve streptopain variants capable of cleaving SpeA by using mRNA display technology. This entirely in vitro method allows the selection of an active enzyme from libraries of up to 1013 variants. We will clone and express SpeA and streptopain, determine the streptopain cleavage site of SmeZ, engineer selection primers displaying the SpeA target region homologous to the SmeZ cleavage site, build a variant streptopain library with random mutations incorporated at specificity conferring regions, and select for SpeA-cleaving ability. We plan to establish the streptopain protease as a platform from which we will develop an arsenal of superantigen-neutralizing proteases to prevent the deadly toxicities of STSS and necrotizing fasciitis as well as defend against the myriad of other diseases linked to superantigens.