Humanization and functional characterization of enhanced coagulation factor IX variants identified through ancestral sequence reconstruction.

ABSTRACT:

Background

Laboratory resurrection of ancient coagulation factor (F) IX variants generated through ancestral sequence reconstruction led to the discovery of a FIX variant, designated An96, which possesses enhanced specific activity independent of and additive to that provided by human p.Arg384Lys, referred to as FIX-Padua.

Objectives

The goal of the current study was to identify the amino acid substitution(s) responsible for the enhanced activity of An96 and create a humanized An96 FIX transgene for gene therapy application.

Methods

Reductionist screening approaches, including domain swapping and scanning residue substitution, were used and guided by one-stage FIX activity assays. In vitro characterization of top candidates included recombinant high-purity preparation

SUBMITTER: Coyle CW 

PROVIDER: S-EPMC10922771 | biostudies-literature | 2024 Mar

REPOSITORIES: biostudies-literature