Strand 1A variant in neuroserpin shows increased aggregation and no loss of inhibition: implication in ameliorating polymerization to retain activity

Biosci Rep. 2022 Dec 22;42(12):BSR20221825. doi: 10.1042/BSR20221825.

Abstract

Neuroserpin (NS) is predominantly expressed in the brain and is the primary inhibitor of tissue plasminogen activator (tPA). NS variants are associated with the neurogenerative disease termed familial encephalopathy with neuroserpin inclusion bodies (FENIB). The disease is characterized by variable age of onset and severity. The reactive center loop (RCL) insertion-based inhibitory mechanism of NS requires a coordinated conformational change leading to a shift in the strands of the β-sheet A and movement of helix F. Strand 1A is connected to the helix F at its C terminal end and with the strand 2A at its N terminal, both these domain move for accommodating the inserting loop; therefore, a variant that influences their movement may alter the inhibition rates. A molecular dynamic simulation analysis of a H138C NS variant from strand 1A showed a large decrease in conformational fluctuations as compared with wild-type NS. H138 was mutated, expressed, purified and a native-PAGE and transmission electron microscopy (TEM) analysis showed that this variant forms large molecular weight aggregates on a slight increase in temperature. However, a circular dichroism analysis showed its secondary structure to be largely conserved. Surprisingly, its tPA inhibition activity and complex formation remain unhindered even after the site-specific labeling of H138C with Alexa fluor C5 maleimide. Further, a helix F-strand 1A (W154C-H138C) double variant still shows appreciable inhibitory activity. Increasingly, it appears that aggregation and not loss of inhibition is the more likely cause of shutter region-based variants phenotypes, indicating that hindering polymer formation using small molecules may retain inhibitory activity in pathological variants of NS.

Keywords: epilepsy; neuroserpin; serpin; tissue plasminogen activator.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Neuropeptides* / genetics
  • Polymerization
  • Serpins* / genetics
  • Tissue Plasminogen Activator

Substances

  • neuroserpin
  • Tissue Plasminogen Activator
  • Serpins
  • Neuropeptides