The Structure of Ca2+ Sensor Case16 Reveals the Mechanism of Reaction to Low Ca2+ Concentrations

被引：12

作者：

Leder, Lukas ^{[2
]}

Stark, Wilhelm ^{[2
,3
]}

Freuler, Felix ^{[2
]}

Marsh, May ^{[2
]}

Meyerhofer, Marco ^{[2
]}

Stettler, Thomas ^{[2
]}

Mayr, Lorenz M. ^{[2
]}

Britanova, Olga V. ^{[1
]}

Strukova, Lydia A. ^{[1
]}

Chudakov, Dmitriy M. ^{[1
]}

Souslova, Ekaterina A. ^{[1
]}

机构：

[1] RAS, Shemiakin Ovchinnikov Inst Bioorgan Chem, Moscow 117997, Russia

[2] Novartis Pharma AG, NIBR CPC LFP, CH-4002 Basel, Switzerland

[3] Fachhsch Nordwestschweiz, Inst Chem & Bioanalyt, CH-4132 Muttenz, Switzerland

来源：

SENSORS | 2010年 / 10卷 / 09期

关键词：

circularly permuted green fluorescent protein; genetically encoded; fluorescent calcium indicator protein; crystal structure; calcium sensor; GREEN FLUORESCENT PROTEINS; IN-VIVO; CALMODULIN; INDICATORS; REFINEMENT; EXPRESSION; PROGRAM; BINDING; GCAMP2;

D O I：

10.3390/s100908143

中图分类号：

O65 [分析化学];

学科分类号：

070302 ; 081704 ;

摘要：

Here we report the first crystal structure of a high-contrast genetically encoded circularly permuted green fluorescent protein (cpGFP)-based Ca2+ sensor, Case16, in the presence of a low Ca2+ concentration. The structure reveals the positioning of the chromophore within Case16 at the first stage of the Ca2+-dependent response when only two out of four Ca2+-binding pockets of calmodulin (CaM) are occupied with Ca2+ ions. In such a "half Ca2+-bound state", Case16 is characterized by an incomplete interaction between its CaM-/M13-domains. We also report the crystal structure of the related Ca2+ sensor Case12 at saturating Ca2+ concentration. Based on this structure, we postulate that cpGFP-based Ca2+ sensors can form non-functional homodimers where the CaM-domain of one sensor molecule binds symmetrically to the M13-peptide of the partner sensor molecule. Case12 and Case16 behavior upon addition of high concentrations of free CaM or M13-peptide reveals that the latter effectively blocks the fluorescent response of the sensor. We speculate that the demonstrated intermolecular interaction with endogenous substrates and homodimerization can impede proper functioning of this type of Ca2+ sensors in living cells.

引用

页码：8143 / 8160

页数：18

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