Structural insights into apoptotic regulation of human Bfk as a novel Bcl-2 family member

Bcl-2 family kin (Bfk), also known as Bcl-2-like 15, plays an essential role in regulating apoptosis by eliciting weak pro-apoptotic responses in the gastrointestinal tract. Human Bfk is a novel Bcl-2 family protein owing to its unique domain composition involving BH2 and BH3. However, the molecular mechanism underlying the regulation of apoptosis by Bfk remains unclear. Here, we first report the crystal structure of human full-length Bfk. Surprisingly, the structure of Bfk adopts a canonical Bcl-2 fold but lacks the hydrophobic cleft, which could accommodate a BH3 domain from other Bcl-2 family proteins. Our biophysical interaction analysis proved that the full-length Bfk itself does not interact with multi-domain Bcl-2 family proteins or a BH3-containing peptide. Instead, Bfk is structurally and functionally reminiscent of Bid, a BH3-only protein in the Bcl-2 family, with similar conformations of helices α3-α5 and the specific motif in helix α5. Not only structural analyses of the full-length Bfk but also molecular dynamics simulation suggested that Bfk elicits its pro-apoptotic activity through a Bid-like apoptotic mechanism in which the BH3 domain is released upon caspase-mediated cleavage and a conformational change of the truncated form. Indeed, the BH3 peptide derived from Bfk exhibited in vitro interactions with Bcl-2, Bcl-X(L), and Bak. These findings provide new insights into the molecular characteristics of Bfk and a valuable foundation for development of a new therapeutic target to control apoptosis.