Although many biosilicification methods based on cationic linear α-poly -l- lysine for synthesis of polylysine/silica hybrids have been investigated, these methods tend to rely on the counteranions, added catalysts, and complex synthesis process. To explore a simple and efficient biosilicification method, in this work, branched poly-l-lysine (BPL) is used as both a catalyst to hydrolyze tetraethoxysilane (TEOS) and an in situ template to direct silicic acids forming polylysine/silica hybrids in one-pot mode. The catalysis of BPL to hydrolyze TEOS results from the abundant hydrogen bonding (as the active site) to increase the nucleophilicity of BPL. Meanwhile, the hydrogen bonding is also found to be the key factor determining the self-assembly of BPL. During biosilicification, owing to self-assembly of BPL molecules, BPL would form spherical particles by keeping a random-coil conformation or form lamellar structures by undergoing a conformational transition from a random-coil to β-sheet construction. As a result, polylysine/silica hybrids with tunable topological structures are synthesized using aggregated BPLs as templates after the hydrolysis of TEOS. This finding of applying BPL to fulfill the biosilicification procedure without counteranions and added catalysts would enable a better understanding of the polypeptide-governed biosilicification process and pave a way for fabricating complex inorganic architectures applicable to silica transformational chemistry.