Bose, K. | bioRxiv Science

bioRxiv2021

Inter-subunit Crosstalk via PDZ Synergistically Governs Allosteric Activation of Proapoptotic HtrA2

Mitochondrial serine protease - High temperature requirement A2 (HtrA2), is associated with various diseases including neurodegenerative disorders and cancer. Despite availability of structural details, the reports on HtrA2s mechanistic regulation that varies with the type of activation signals still remain non-concordant. To expound the role of regulatory PDZ domains in promoting synergistic coordination between HtrA2 subunits, we generated heterotrimeric HtrA2 variants comprising different numbers of PDZs and/or active-site mutations. Sequential deletion of PDZs from the trimeric ensemble significantly affected its residual activity in a way that proffered a hypothesis advocating intermolecular allosteric crosstalk via PDZ domains in trimeric HtrA2. Furthermore, structural and computational snapshots affirmed the role of PDZs in secondary structural element formation and coordinated reorganization of the N-terminal region and regulatory loops. Therefore, apart from providing cues for devising structure-guided therapeutic strategies, this study establishes a physiologically relevant working model of complex allosteric regulation through a multifaceted trans-mediated cooperatively-shared energy landscape.

biochemistry↗

bioRxiv2021

Simultaneous Control of Infection and Inflammation by Keratin-Derived Antimicrobial Peptides (KAMPs) Targeting TLRs and Co-receptors

The use and the timing of initiation of steroids for controlling unwanted infectious inflammation are major clinical dilemmas due to their possible adverse effects including delayed microbial clearance and wound healing. Compounding this difficulty is the continued emergence of drug-resistant bacteria; yet anti-infective strategies aiming at augmenting inflammatory responses to facilitate bacterial killing cannot be used to treat infections involving vulnerable tissues. As is the case with bacterial keratitis, excessive inflammation jeopardizes corneal transparency leading to devastating vision loss. Hence, a two-pronged remedy possessing both anti-infective and anti-inflammatory properties would be helpful for tackling antibiotic resistance and enabling prompt inflammation control at once. Using murine primary neutrophils, macrophages and sterile corneal inflammation models, we found that non-toxic and pro-healing human keratin 6a-derived antimicrobial peptides (KAMPs) with a native 10-or 18-amino-acid sequence suppress LTA- and LPS-induced NF-[kcy]B and IRF3 activation, proinflammatory cytokine production, as well as phagocyte recruitment, independently of their bactericidal function. Mechanistically, direct binding of KAMPs to cell surface TLR2 and TLR co-receptors CD14 and MD-2 not only blocks their bacterial ligand docking sites, but also reduces cell surface availability of TLR2 and TLR4 through promotion of receptor endocytosis. Benefitting from the dual functions of topical KAMPs, experimental bacterial keratitis caused was effectively prevented or controlled, as evidenced by significant reductions of corneal opacification and inflammatory cell infiltration in addition to enhanced bacterial clearance. These findings reveal multiple TLR-targeting activities of KAMPs and demonstrate their therapeutic potential as a multifunctional drug for managing sterile and infectious inflammatory diseases. One Sentence SummaryBifunctional native keratin peptides allow concurrent alleviation of inflammation and infection to avoid functional damages in vulnerable tissues.

immunology↗