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Structural insights into the unique inhibitory mechanism of the silkworm protease inhibitor serpin18.

Guo PC, Dong Z, Zhao P, Zhang Y, He H, Tan X, Zhang W, Xia Q - Sci Rep (2015)

Bottom Line: Notably, this inhibitiory reaction results from the formation of an intermediate complex, which then follows for the digestion of protease and inhibitor into small fragments.This activity differs from previously reported modes of inhibition for serpins.Our findings have thus provided novel structural insights into the unique inhibitory mechanism of serpin18.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, 216, Tiansheng Road, Beibei, Chongqing 400716, People's Republic of China.

ABSTRACT
Serpins generally serve as inhibitors that utilize a mobile reactive center loop (RCL) as bait to trap protease targets. Here, we present the crystal structure of serpin18 from Bombyx mori at 1.65 Å resolution, which has a very short and stable RCL. Activity analysis showed that the inhibitory target of serpin18 is a cysteine protease rather than a serine protease. Notably, this inhibitiory reaction results from the formation of an intermediate complex, which then follows for the digestion of protease and inhibitor into small fragments. This activity differs from previously reported modes of inhibition for serpins. Our findings have thus provided novel structural insights into the unique inhibitory mechanism of serpin18. Furthermore, one physiological target of serpin18, fibroinase, was identified, which enables us to better define the potential role for serpin18 in regulating fibroinase activity during B. mori development.

No MeSH data available.


Related in: MedlinePlus

Overall structure of serpin18.(A) Schematic representation of serpin18 fold. Blue, helical region; red, RCL; and the three β-sheets are distinguished by diffferent colour (sA, orange; sB, yellow; sC, magenta). Secondary elements are labeled, and the specific numbering of strands 1-3, 5, and 6 of sA is shown. (B) The residues in RCL are emphasized in stick presentation with the corresponding electron density at a sigma level of 1.0 in the 2Fo-Fc map. (C) Residues involved the interactions between the center of RCL and sC are shown with sticks, and the hydrogen bonds are black dashed lines. All figures were prepared using PyMOL.
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f1: Overall structure of serpin18.(A) Schematic representation of serpin18 fold. Blue, helical region; red, RCL; and the three β-sheets are distinguished by diffferent colour (sA, orange; sB, yellow; sC, magenta). Secondary elements are labeled, and the specific numbering of strands 1-3, 5, and 6 of sA is shown. (B) The residues in RCL are emphasized in stick presentation with the corresponding electron density at a sigma level of 1.0 in the 2Fo-Fc map. (C) Residues involved the interactions between the center of RCL and sC are shown with sticks, and the hydrogen bonds are black dashed lines. All figures were prepared using PyMOL.

Mentions: The overall fold of serpin18 resembles the canonical serpin conformation: an α/β fold consisting of one core β-sheets surrounded by nine α-helices (hA–hI) that are classified according to the conventional serpin nomenclature22. The core β-sheets includes three antiparallel β-sheets (sA: β1, β2, β3, β5, and β6; sB; and sC) (Fig. 1A and Fig. S1). Additionally, one RCL, which functions as the bait for the target protease in classical serpins, is solvent-exposed and located on the top of the molecule. Specifically, the segment with residues Glu352–Pro368 makes up the active RCL region, which is folded into a 1.5-turn helix (Glu355–Ser360) and two defined loops (Gln352–Thr354 and Gln361–Pro365, respectively). All residues of the RCL were well defined in the final electron density map (Fig. 1B), presumably because of the interaction between the RCL region and the underlying protein scaffold. Notably, the carbonyl oxygens of Val361 and Ala358 make two hydrogen bonds with Nζ of Lys206. The amide nitrogen of Glu364 forms a hydrogen bond with the hydroxyl group of Tyr226. Moreover, hydrophobic contacts between the aliphatic side chains of Val361 and Tyr205, with Ala361 and Tyr226, respectively, also contribute to the stable conformation of RCL (Fig. 1C).


Structural insights into the unique inhibitory mechanism of the silkworm protease inhibitor serpin18.

Guo PC, Dong Z, Zhao P, Zhang Y, He H, Tan X, Zhang W, Xia Q - Sci Rep (2015)

Overall structure of serpin18.(A) Schematic representation of serpin18 fold. Blue, helical region; red, RCL; and the three β-sheets are distinguished by diffferent colour (sA, orange; sB, yellow; sC, magenta). Secondary elements are labeled, and the specific numbering of strands 1-3, 5, and 6 of sA is shown. (B) The residues in RCL are emphasized in stick presentation with the corresponding electron density at a sigma level of 1.0 in the 2Fo-Fc map. (C) Residues involved the interactions between the center of RCL and sC are shown with sticks, and the hydrogen bonds are black dashed lines. All figures were prepared using PyMOL.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493575&req=5

f1: Overall structure of serpin18.(A) Schematic representation of serpin18 fold. Blue, helical region; red, RCL; and the three β-sheets are distinguished by diffferent colour (sA, orange; sB, yellow; sC, magenta). Secondary elements are labeled, and the specific numbering of strands 1-3, 5, and 6 of sA is shown. (B) The residues in RCL are emphasized in stick presentation with the corresponding electron density at a sigma level of 1.0 in the 2Fo-Fc map. (C) Residues involved the interactions between the center of RCL and sC are shown with sticks, and the hydrogen bonds are black dashed lines. All figures were prepared using PyMOL.
Mentions: The overall fold of serpin18 resembles the canonical serpin conformation: an α/β fold consisting of one core β-sheets surrounded by nine α-helices (hA–hI) that are classified according to the conventional serpin nomenclature22. The core β-sheets includes three antiparallel β-sheets (sA: β1, β2, β3, β5, and β6; sB; and sC) (Fig. 1A and Fig. S1). Additionally, one RCL, which functions as the bait for the target protease in classical serpins, is solvent-exposed and located on the top of the molecule. Specifically, the segment with residues Glu352–Pro368 makes up the active RCL region, which is folded into a 1.5-turn helix (Glu355–Ser360) and two defined loops (Gln352–Thr354 and Gln361–Pro365, respectively). All residues of the RCL were well defined in the final electron density map (Fig. 1B), presumably because of the interaction between the RCL region and the underlying protein scaffold. Notably, the carbonyl oxygens of Val361 and Ala358 make two hydrogen bonds with Nζ of Lys206. The amide nitrogen of Glu364 forms a hydrogen bond with the hydroxyl group of Tyr226. Moreover, hydrophobic contacts between the aliphatic side chains of Val361 and Tyr205, with Ala361 and Tyr226, respectively, also contribute to the stable conformation of RCL (Fig. 1C).

Bottom Line: Notably, this inhibitiory reaction results from the formation of an intermediate complex, which then follows for the digestion of protease and inhibitor into small fragments.This activity differs from previously reported modes of inhibition for serpins.Our findings have thus provided novel structural insights into the unique inhibitory mechanism of serpin18.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, 216, Tiansheng Road, Beibei, Chongqing 400716, People's Republic of China.

ABSTRACT
Serpins generally serve as inhibitors that utilize a mobile reactive center loop (RCL) as bait to trap protease targets. Here, we present the crystal structure of serpin18 from Bombyx mori at 1.65 Å resolution, which has a very short and stable RCL. Activity analysis showed that the inhibitory target of serpin18 is a cysteine protease rather than a serine protease. Notably, this inhibitiory reaction results from the formation of an intermediate complex, which then follows for the digestion of protease and inhibitor into small fragments. This activity differs from previously reported modes of inhibition for serpins. Our findings have thus provided novel structural insights into the unique inhibitory mechanism of serpin18. Furthermore, one physiological target of serpin18, fibroinase, was identified, which enables us to better define the potential role for serpin18 in regulating fibroinase activity during B. mori development.

No MeSH data available.


Related in: MedlinePlus