title: Penta-L-lysine Potentiates Fibrin-Independent Activity of Human Tissue Plasminogen Activator.
abstract: The therapeutic action of tissue plasminogen activator (t-PA) is a two-step process: (1) binding to lysine-rich fibrin (Km event) and (2) converting local plasminogen into plasmin (Kcat event). Overcoming limitations of other structural biophysics methods, we wanted to employ small-angle X-ray scattering (SAXS) to visualize what shape changes occur/accompany t-PA activation, but the prime hurdle was the polydisperse nature of the fibrin, which occluded scattering information from t-PA. Earlier, larger polylysine peptides have been used to potentiate activation of t-PA, so while screening short polylysine peptides as alternatives to fibrin or larger peptides, we found that penta-polylysine (P5) specifically activates t-PA in a dose-dependent manner, averaging to almost 3-fold more than in the absence of any peptide. SAXS data analysis confirmed that P5 does not induce association of t-PA molecules, and a narrower peak profile of the Kratky plot indicated that P5 binding quenches inherent motion in t-PA. Shape reconstruction of t-PA ∓ P5 revealed that P5 closes the "gap" between the two gross domains of t-PA, viz. fused F/E, K1 and K2 domains, and the P domain. Docking experiments suggested that, while other polylysine peptides preferentially interacted with the surfaces of kringle domains, P5 "slipped into" the gap/groove between K2 and P domains, thereby mediating a substantial increase in the number of long-range interactions between the K2 domain and exosites in the P domain. We report here dissection of shape events involved in between Km/Kcat steps of t-PA activation, which can pave the way toward the search for small molecule function regulator(s) of t-PA.
authors.affiliation: CSIR-Institute of Microbial Technology , Sector 39-A, Chandigarh, 160036, India.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Rehan
authors.affiliation: CSIR-Institute of Microbial Technology , Sector 39-A, Chandigarh, 160036, India.
authors.forename: Amin
authors.initials: A
authors.lastname: Sagar
authors.affiliation: CSIR-Institute of Microbial Technology , Sector 39-A, Chandigarh, 160036, India.
authors.forename: Vandna
authors.initials: V
authors.lastname: Sharma
authors.affiliation: CSIR-Institute of Microbial Technology , Sector 39-A, Chandigarh, 160036, India.
authors.forename: Sanskruti
authors.initials: S
authors.lastname: Mishra
authors.affiliation: CSIR-Institute of Microbial Technology , Sector 39-A, Chandigarh, 160036, India.
authors.lastname: Ashish
authors.affiliation: CSIR-Institute of Microbial Technology , Sector 39-A, Chandigarh, 160036, India.
authors.forename: Girish
authors.initials: G
authors.lastname: Sahni
mesh_terms: D005337:Fibrin / Q000378:metabolism*; D006801:Humans; D062105:Molecular Docking Simulation; D009842:Oligopeptides / Q000494:pharmacology*; D010959:Tissue Plasminogen Activator / Q000378:metabolism*
chemical_list: D009842:Oligopeptides; C026591:pentalysine; D005337:Fibrin; D010959:Tissue Plasminogen Activator