Probing intermolecular backbone H-bonding in serine proteinase-protein inhibitor complexes
Research article published in Chemistry & biology (1999)
Abstract
BACKGROUND: Intermolecular backbone H-bonding (N-H.O=C) is a common occurrence at the interface of protein-protein complexes. For instance, the amide NH groups of most residues in the binding loop of eglin c, a potent serine proteinase inhibitor from the leech Hirudo medicinalis, are H-bonded to the carbonyl groups of residues in the target enzyme molecules such as chymotrypsin, elastase and subtilisins. We sought to understand the energetic significance of these highly conserved backbone-backbone H-bonds in the enzyme-inhibitor complexes. RESULTS: We synthesized an array of backbone-engineered ester analogs of eglin c using native chemical ligation to yield five inhibitor proteins each containing a single backbone ester bond from P3 to P2' (i.e. -CONH-to -COO-). The structure at the ligation site (P6-P5) is essentially unaltered as shown by a high-resolution analysis of the subtilisin-BPN'-eglin c complex. The free-energy changes (DeltaDeltaGNH-->O) associated with the binding of ester analogs at P3, P1 and P2' with bovine alpha-chymotrypsin, subtilisin Carlsberg and porcine pancreatic elastase range from 0-4.5 kcal/mol. Most markedly, the NH-->O substitution at P2 not only stabilizes the inhibitor but also enhances binding to the enzymes by as much as 500-fold. CONCLUSIONS: Backbone H-bond contributions are context dependent in the enzyme-eglin c complexes. The interplay of rigidity and adaptability of the binding loop of eglin c seems to play a prominent role in defining the binding action.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Resumen
Intermolecular backbone H-bonding (N-H.O=C) is a common occurrence at the interface of protein-protein complexes.
Por qué esto importa para la hirudoterapia
Este estudio empleó la ligación química nativa para sintetizar análogos éster del eglin c con backbone modificado —un potente inhibidor de serina proteinasas de la sanguijuela Hirudo medicinalis— con el fin de sondear las contribuciones energéticas de los enlaces de hidrógeno intermoleculares del backbone en la interfaz enzima-inhibidor. Se ensayaron cinco análogos éster con modificaciones puntuales del backbone desde P3 hasta P2' frente a α-quimotripsina bovina, subtilisina Carlsberg y elastasa pancreática porcina, revelando cambios de energía libre dependientes del contexto (0–4,5 kcal/mol) y un notable aumento de 500 veces en la unión a partir de una sola sustitución NH→O en P2. Este trabajo es directamente relevante para el ámbito de ASH, ya que aporta conocimiento detallado a nivel molecular sobre el mecanismo de unión de un inhibidor de proteinasas derivado de sanguijuela. La matización es que se trata de un estudio puramente in vitro de química estructural y biofísica, sin implicaciones terapéuticas ni clínicas abordadas directamente; los hallazgos conciernen a la energética básica de las interacciones proteína-proteína y no a aplicaciones de hirudoterapia.
Citación
Probing intermolecular backbone H-bonding in serine proteinase-protein inhibitor complexes
Lu W et al. · Chemistry & biology, 1999
Contexto clínico relacionado
Explore cómo esta investigación se conecta con la práctica clínica
Añadido a la biblioteca ASH: May 27, 2026 · Última actualización del sitio: 18 de junio de 2026