title: Immobilized thermolysin for highly efficient production of low-molecular-weight protamine--an attractive cell-penetrating peptide for macromolecular drug delivery applications.
abstract: Macromolecules present a remarkable potential as future therapeutics. However, their translation into clinical practice has been hampered by an inherently low bioavailability. Cell-penetrating peptides (CPP) have been recently shown to significantly improve on the bioavailability of macromolecules. Yet, the high cost associated with development and production of these peptides is a major factor hindering their rapid deployment beyond the laboratory. Here, we describe a facile and robust methodology for efficient and large-scale production of low-molecular-weight protamine-a potent CPP of great clinical potential. Our methodology is based on the immobilization of thermolysin, an enzyme catalyzing digestion of native protamine, on chemically surface-modified gels produced by silica sol-gel chemistry. Thermolysin was immobilized at extremely high matrix loading of 733 mg/g matrix and exhibited good thermal and pH stability, indicating robustness with respect to processing conditions. The mechanical properties of the silica matrix further allowed utilization of the immobilized thermolysin in both batch and packed-bed reactor systems to produce the LMWP peptide in high yields. Results presented here are of high significance as this efficient and cost-effective production of high purity LMWP could enable clinical translation of many potential macromolecular drugs.
authors: Department of Chemical Engineering, A. James Clark School of Engineering, University of Maryland, College Park, Maryland 20742, USA.
authors: Allan E
authors: AE
authors: David
authors: Junbo
authors: J
authors: Gong
authors: Beata
authors: B
authors: Chertok
authors: Roman C
authors: RC
authors: Domszy
authors: Cheol
authors: C
authors: Moon
authors: Yoon Shin
authors: YS
authors: Park
authors: Nam Sun
authors: NS
authors: Wang
authors: Arthur J
authors: AJ
authors: Yang
authors: Victor C
authors: VC
authors: Yang
mesh_terms: D001407:Bacillus / Q000201:enzymology; D019149:Bioreactors; D057846:Cell-Penetrating Peptides / Q000096:biosynthesis* / Q000737:chemistry / Q000494:pharmacology; D002846:Chromatography, Affinity; D016503:Drug Delivery Systems / Q000379:methods*; D004800:Enzymes, Immobilized / Q000378:metabolism*; D005782:Gels; D005976:Glutaral / Q000737:chemistry; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007700:Kinetics; D046911:Macromolecular Substances / Q000378:metabolism*; D008855:Microscopy, Electron, Scanning; D008970:Molecular Weight; D044367:Phase Transition / Q000187:drug effects; D016062:Porosity / Q000187:drug effects; D011479:Protamines / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D012822:Silicon Dioxide / Q000737:chemistry; D013499:Surface Properties / Q000187:drug effects; D013696:Temperature; D013820:Thermolysin / Q000378:metabolism*
chemical_list: D057846:Cell-Penetrating Peptides; D004800:Enzymes, Immobilized; D005782:Gels; D046911:Macromolecular Substances; D011479:Protamines; D012822:Silicon Dioxide; D013820:Thermolysin; D005976:Glutaral