Dataset Information

Ca2+ activation of the cPLA2 C2 domain: ordered binding of two Ca2+ ions with positive cooperativity.

ABSTRACT: During Ca(2+) activation, the Ca(2+)-binding sites of C2 domains typically bind multiple Ca(2+) ions in close proximity. These binding events exhibit positive cooperativity, despite the strong charge repulsion between the adjacent divalent cations. Using both experimental and computational approaches, the present study probes the detailed mechanisms of Ca(2+) activation and positive cooperativity for the C2 domain of cytosolic phospholipase A(2), which binds two Ca(2+) ions in sites I and II, separated by only 4.1 A. First, each of the five coordinating side chains in the Ca(2+)-binding cleft is individually mutated and the effect on Ca(2+)-binding affinity and cooperativity is measured. The results identify Asp 43 as the major contributor to Ca(2+) affinity, while the two coordinating side chains that provide bridging coordination to both Ca(2+) ions, Asp 43 and Asp 40, are observed to make the largest contributions to positive cooperativity. Electrostatic calculations reveal that Asp 43 possesses the highest pseudo-pK(a) of the coordinating acidic residues, as well as the highest general cation affinity, due to its relatively buried location within 3.5 A of seven protein oxygens with full or partial negative charges. These calculations therefore explain the greater importance of Asp 43 in defining the Ca(2+) affinity. Overall, the experimental and computational results support an activation model in which the first Ca(2+) ion binds usually to site I, thereby preordering both bridging side chains Asp 40 and 43, and partially or fully deprotonating the three coordinating Asp residues. This initial binding event prepares the conformation and protonation state of the remaining site for Ca(2+) binding, enabling the second Ca(2+) ion to bind with higher affinity than the first as required for positive cooperativity.

SUBMITTER: Malmberg NJ

PROVIDER: S-EPMC3657617 | biostudies-literature | 2004 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Ca2+ activation of the cPLA2 C2 domain: ordered binding of two Ca2+ ions with positive cooperativity.

Malmberg Nathan J NJ Varma Sameer S Jakobsson Eric E Falke Joseph J JJ

Biochemistry 20041201 51

During Ca(2+) activation, the Ca(2+)-binding sites of C2 domains typically bind multiple Ca(2+) ions in close proximity. These binding events exhibit positive cooperativity, despite the strong charge repulsion between the adjacent divalent cations. Using both experimental and computational approaches, the present study probes the detailed mechanisms of Ca(2+) activation and positive cooperativity for the C2 domain of cytosolic phospholipase A(2), which binds two Ca(2+) ions in sites I and II, se ...[more]

PMID: 15610026

Similar Datasets

Project description:Temperature is considered one of the critical factors directly influencing grapevine during the three primary growth and development stages: sprout, flowering, and fruit-coloring, which is strongly correlated to the yield and quality of the grape. The grapevine is frequently exposed to high-temperature conditions that are detrimental to growth. However, the mechanisms of the heat stress response and adaptation in grapevine are not adequately studied. The Arabidopsis copine gene AtBON1 encodes a highly conserved protein containing two C2 domains at the amino terminus, participation in cell death regulation and defense responses. Previously, we showed that a BON1 association protein from the grapevine, VvBAP1, plays a positive role in cold tolerance. Similarly, the involvement of VvBAP1 in the resistance to heat stress was also found in the present study. The results indicated VvBAP1 was significantly induced by high temperature, and the elevated expression of VvBAP1 was significantly higher in the resistant cultivars than the sensitive cultivars under heat stress. Seed germination and phenotypic analysis results indicated that overexpression of VvBAP1 improved Arabidopsis thermoresistance. Compared with the wild type, the chlorophyll content and net photosynthetic rate in VvBAP1 overexpressing Arabidopsis plants were markedly increased under heat stress. At high temperatures, overexpression of VvBAP1 also enhanced antioxidant enzyme activity as well as their corresponding gene transcription levels, to reduce the accumulation of reactive oxygen species and lipid peroxidation. Besides, the transcriptional activities of HSP70, HSP101, HSFA2, and HSFB1 in VvBAP1 overexpressing Arabidopsis plants were significantly up-regulated compare to the wild type. In summary, we propose that VvBAP1 may play a potential important role in enhanced grapevine thermoresistance, primarily through the enhancement of antioxidant enzyme activity and promoted heat stress response genes expression.

Dataset Information

Ca2+ activation of the cPLA2 C2 domain: ordered binding of two Ca2+ ions with positive cooperativity.

Publications

Ca2+ activation of the cPLA2 C2 domain: ordered binding of two Ca2+ ions with positive cooperativity.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets