The switch from client holding to folding in the Hsp70/Hsp90 chaperone machineries is regulated by a direct interplay between co-chaperones.

Folding of stringent clients requires transfer from Hsp70 to Hsp90. The co-chaperone Hop physically connects the chaperone machineries. Here, we define its role from the remodeling of Hsp70/40-client complexes to the mechanism of client transfer and the conformational switching from stalled to active client-processing states of Hsp90. We show that Hsp70 together with Hsp40 completely unfold a stringent client, the glucocorticoid receptor ligand-binding domain (GR-LBD) in large assemblies. Hop remodels these for efficient transfer onto Hsp90. As p23 enters, Hsp70 leaves the complex via switching between binding sites in Hop. Current concepts assume that to proceed to client folding, Hop dissociates and the co-chaperone p23 stabilizes the Hsp90 closed state. In contrast, we show that p23 functionally interacts with Hop, relieves the stalling Hsp90-Hop interaction, and closes Hsp90. This reaction allows folding of the client and is thus the key regulatory step for the progression of the chaperone cycle.

Kindlin stabilizes the talin·integrin bond under mechanical load by generating an ideal bond.

Integrin-mediated adhesion is essential for metazoan life. Integrin binding to ligand requires an activation step prior to binding ligand that depends on direct binding of talin and kindlin to the ß-integrin cytoplasmic tail and the transmission of force from the actomyosin via talin to the integrin-ligand bonds. However, the affinity of talin for integrin tails is low. It is therefore still unclear how such low-affinity bonds are reinforced to transmit forces up to 10 to 40 pN. In this study, we use single-molecule force spectroscopy by optical tweezers to investigate the mechanical stability of the talin•integrin bond in the presence and absence of kindlin. While talin and integrin alone form a weak and highly dynamic slip bond, the addition of kindlin-2 induces a force-independent, ideal talin•integrin bond, which relies on the steric proximity of and the intervening amino acid sequences between the talin- and kindlin-binding sites in the ß-integrin tail. Our findings show how kindlin cooperates with talin to enable transmission of high forces required to stabilize cell adhesion.

Active unfolding of the glucocorticoid receptor by the Hsp70/Hsp40 chaperone system in single-molecule mechanical experiments.

The glucocorticoid receptor (GR) is an important transcription factor and drug target linked to a variety of biological functions and diseases. It is one of the most stringent physiological clients of the Hsp90/Hsp70/Hsp40 chaperone system. In this study, we used single-molecule force spectroscopy by optical tweezers to observe the interaction of the GR's ligand-binding domain (GR-LBD) with the Hsp70/Hsp40 chaperone system (Hsp70/40). We show in real time that Hsp70/40 can unfold the complete GR-LBD in a stepwise manner. Each unfolding step involves binding of an Hsp70 to the GR-LBD and subsequent adenosine triphosphate (ATP) hydrolysis, stimulated by Hsp40. The kinetics of chaperone-mediated unfolding depend on chaperone concentrations as well as the presence of the nucleotide exchange factor BAG1. We find that Hsp70/40 can stabilize new unfolding intermediates, showing that Hsp70/40 can directly interact with the folded core of the protein when working as an unfoldase. Our results support an unfolding mechanism where Hsp70 can directly bind to folded protein structures and unfold them upon ATP hydrolysis. These results provide important insights into the regulation of GR by Hsp70/40.

Direct observation of chemo-mechanical coupling in DnaK by single-molecule force experiments.

Protein allostery requires a communication channel for functional regulation between distal sites within a protein. In the molecular chaperone Hsp70, a two-domain enzyme, the ATP/ADP status of an N-terminal nucleotide-binding domain regulates the substrate affinity of a C-terminal substrate-binding domain. Recently available three-dimensional structures of Hsp70 in ATP/ADP states have provided deep insights into molecular pathways of allosteric signals. However, direct mechanical probing of long-range allosteric coupling between the ATP hydrolysis step and domain states is missing. Using laser optical tweezers, we examined the mechanical properties of a truncated two-domain DnaK(1-552ye) in apo/ADP/ATP- and peptide-bound states. We find that in the apo and ADP states, DnaK domains are mechanically stable and rigid. However, in the ATP state, substrate-binding domain (SBD)∗ye is mechanically destabilized as the result of interdomain docking followed by the unfolding of the α-helical lid. By observing the folding state of the SBD, we could observe the continuous ATP/ADP cycling of the enzyme in real time with a single molecule. The SBD lid closure is strictly coupled to the chemical steps of the ATP hydrolysis cycle even in the presence of peptide substrate.

Detection of relevant extracardiac findings on coronary computed tomography angiography vs. invasive coronary angiography.

OBJECTIVES: To compare the detection of relevant extracardiac findings (ECFs) on coronary computed tomography angiography (CTA) and invasive coronary angiography (ICA) and evaluate the potential clinical benefit of their detection. METHODS: This is the prespecified subanalysis of ECFs in patients presenting with a clinical indication for ICA based on atypical angina and suspected coronary artery disease (CAD) included in the prospective single-center randomized controlled Coronary Artery Disease Management (CAD-Man) study. ECFs requiring immediate therapy and/or further workup including additional imaging were defined as clinically relevant. We evaluated the scope of ECFs in 329 patients and analyzed the potential clinical benefit of their detection. RESULTS: ECFs were detected in 107 of 329 patients (32.5%; CTA: 101/167, 60.5%; ICA: 6/162, 3.7%; p < .001). Fifty-nine patients had clinically relevant ECFs (17.9%; CTA: 55/167, 32.9%; ICA: 4/162, 2.5%; p < .001). In the CTA group, ECFs potentially explained atypical chest pain in 13 of 101 patients with ECFs (12.9%). After initiation of therapy, chest pain improved in 4 (4.0%) and resolved in 7 patients (6.9%). Follow-up imaging was recommended in 33 (10.0%; CTA: 30/167, 18.0%; ICA: 3/162, 1.9%) and additional clinic consultation in 26 patients (7.9%; CTA: 25/167, 15.0%; ICA: 1/162, 0.6%). Malignancy was newly diagnosed in one patient (0.3%; CTA: 1/167, 0.6%; ICA: 0). CONCLUSIONS: In this randomized study, CTA but not ICA detected clinically relevant ECFs that may point to possible other causes of chest pain in patients without CAD. Thus, CTA might preclude the need for ICA in those patients. TRIAL REGISTRATION: NCT Unique ID: 00844220 KEY POINTS: • CTA detects ten times more clinically relevant ECFs than ICA. • Actionable clinically relevant ECFs affect patient management and therapy and may thus improve chest pain. • Detection of ECFs explaining chest pain on CTA might preclude the need for performing ICA.

Patient preferences for development in MRI scanner design: a survey of claustrophobic patients in a randomized study.

OBJECTIVE: To investigate which magnetic resonance imaging (MRI) scanner designs claustrophobic patients prefer. MATERIAL/METHODS: We analyzed questionnaires completed by 160 patients at high risk for claustrophobia directly after a scan in either a short-bore or open panoramic scanner as part of a prospective randomized trial Enders et al (BMC Med Imaging 11:4, 2011). Scanner preferences were judged based on schematic drawings of four scanners. Information on the diagnostic performance of the depicted scanners was provided, too. RESULTS: A majority of patients suggested upright open (59/160, 36.9%) and open panoramic (53/160, 33.1%) before short-bore designs (26/160, 16.3%, for all p < 0.001) for future development. When asked about patients' preferred scanner choice for an upcoming examination, information about a better diagnostic performance of a short-bore scanner significantly improved its preference rates (from 6/160 to 49/160 or 3.8 to 30.5%, p < 0.001). Patients with a claustrophobic event preferred open designs significantly more often than patients without a claustrophobic event (p = 0.047). Patients scanned in a short-bore scanner in our trial preferred this design significantly more often (p = 0.003). Noise reduction (51/160, 31.9%), more space over the head (44/160, 27.5%), and overall more space (33/160, 20.6%) were the commonest suggested areas of improvement. CONCLUSION: Patients at high risk for claustrophobia visually prefer open- over short-bore MRI designs for further development. Education about a better diagnostic performance of a visually less-attractive scanner can increase its acceptance. Noise and space were of most concern for claustrophobic patients. This information can guide individual referral of claustrophobic patients to scanners and future scanner development. KEY POINTS: • Patients at high risk for claustrophobia visually favor the further development of open scanners as opposed to short- and closed-bore scanner designs. • Educating claustrophobic patients about a higher diagnostic performance of a short-bore scanner can significantly increase their acceptance of this otherwise visually less-attractive design. • A medical history of earlier claustrophobic events in a given MRI scanner type and focusing on the features "more space" and "noise reduction" can help to guide referral of patients who are at high risk for claustrophobia.

Single-molecule force spectroscopy reveals folding steps associated with hormone binding and activation of the glucocorticoid receptor.

The glucocorticoid receptor (GR) is a prominent nuclear receptor linked to a variety of diseases and an important drug target. Binding of hormone to its ligand binding domain (GR-LBD) is the key activation step to induce signaling. This process is tightly regulated by the molecular chaperones Hsp70 and Hsp90 in vivo. Despite its importance, little is known about GR-LBD folding, the ligand binding pathway, or the requirement for chaperone regulation. In this study, we have used single-molecule force spectroscopy by optical tweezers to unravel the dynamics of the complete pathway of folding and hormone binding of GR-LBD. We identified a "lid" structure whose opening and closing is tightly coupled to hormone binding. This lid is located at the N terminus without direct contacts to the hormone. Under mechanical load, apo-GR-LBD folds stably and readily without the need of chaperones with a folding free energy of [Formula: see text] The folding pathway is largely independent of the presence of hormone. Hormone binds only in the last step and lid closure adds an additional [Formula: see text] of free energy, drastically increasing the affinity. However, mechanical double-jump experiments reveal that, at zero force, GR-LBD folding is severely hampered by misfolding, slowing it to less than 1·s-1 From the force dependence of the folding rates, we conclude that the misfolding occurs late in the folding pathway. These features are important cornerstones for understanding GR activation and its tight regulation by chaperones.

A folding nucleus and minimal ATP binding domain of Hsp70 identified by single-molecule force spectroscopy.

The folding pathways of large proteins are complex, with many of them requiring the aid of chaperones and others folding spontaneously. Along the folding pathways, partially folded intermediates are frequently populated; their role in the driving of the folding process is unclear. The structures of these intermediates are generally not amenable to high-resolution structural techniques because of their transient nature. Here we employed single-molecule force measurements to scrutinize the hierarchy of intermediate structures along the folding pathway of the nucleotide binding domain (NBD) of Escherichia coli Hsp70 DnaK. DnaK-NBD is a member of the sugar kinase superfamily that includes Hsp70s and the cytoskeletal protein actin. Using optical tweezers, a stable nucleotide-binding competent en route folding intermediate comprising lobe II residues (183-383) was identified as a critical checkpoint for productive folding. We obtained a structural snapshot of this folding intermediate that shows native-like conformation. To assess the fundamental role of folded lobe II for efficient folding, we turned our attention to yeast mitochondrial NBD, which does not fold without a dedicated chaperone. After replacing the yeast lobe II residues with stable E. coli lobe II, the obtained chimeric protein showed native-like ATPase activity and robust folding into the native state, even in the absence of chaperone. In summary, lobe II is a stable nucleotide-binding competent folding nucleus that is the key to time-efficient folding and possibly resembles a common ancestor domain. Our findings provide a conceptual framework for the folding pathways of other members of this protein superfamily.

Multiplexing molecular tension sensors reveals piconewton force gradient across talin-1.

Förster resonance energy transfer (FRET)-based tension sensor modules (TSMs) are available for investigating how distinct proteins bear mechanical forces in cells. Yet, forces in the single piconewton (pN) regime remain difficult to resolve, and tools for multiplexed tension sensing are lacking. Here, we report the generation and calibration of a genetically encoded, FRET-based biosensor called FL-TSM, which is characterized by a near-digital force response and increased sensitivity at 3-5 pN. In addition, we present a method allowing the simultaneous evaluation of coexpressed tension sensor constructs using two-color fluorescence lifetime microscopy. Finally, we introduce a procedure to calculate the fraction of mechanically engaged molecules within cells. Application of these techniques to new talin biosensors reveals an intramolecular tension gradient across talin-1 that is established upon integrin-mediated cell adhesion. The tension gradient is actomyosin- and vinculin-dependent and sensitive to the rigidity of the extracellular environment.

Torque generation of kinesin motors is governed by the stability of the neck domain.

In long-range transport of cargo, prototypical kinesin-1 steps along a single protofilament on the microtubule, an astonishing behavior given the number of theoretically available binding sites on adjacent protofilaments. Using a laser trap assay, we analyzed the trajectories of several representatives from the kinesin-2 class on freely suspended microtubules. In stark contrast to kinesin-1, these motors display a wide range of left-handed spiraling around microtubules and thus generate torque during cargo transport. We provide direct evidence that kinesin's neck region determines the torque-generating properties. A model system based on kinesin-1 corroborates this result: disrupting the stability of the neck by inserting flexible peptide stretches resulted in pronounced left-handed spiraling. Mimicking neck stability by crosslinking significantly reduced the spiraling of the motor up to the point of protofilament tracking. Finally, we present a model that explains the physical basis of kinesin's spiraling around the microtubule.

Mechanically switching single-molecule fluorescence of GFP by unfolding and refolding.

Green fluorescent protein (GFP) variants are widely used as genetically encoded fluorescent fusion tags, and there is an increasing interest in engineering their structure to develop in vivo optical sensors, such as for optogenetics and force transduction. Ensemble experiments have shown that the fluorescence of GFP is quenched upon denaturation. Here we study the dependence of fluorescence on protein structure by driving single molecules of GFP into different conformational states with optical tweezers and simultaneously probing the chromophore with fluorescence. Our results show that fluorescence is lost during the earliest events in unfolding, 3.5 ms before secondary structure is disrupted. No fluorescence is observed from the unfolding intermediates or the ensemble of compact and extended states populated during refolding. We further demonstrate that GFP can be mechanically switched between emissive and dark states. These data definitively establish that complete structural integrity is necessary to observe single-molecule fluorescence of GFP.

α-Actinin/titin interaction: A dynamic and mechanically stable cluster of bonds in the muscle Z-disk.

Stable anchoring of titin within the muscle Z-disk is essential for preserving muscle integrity during passive stretching. One of the main candidates for anchoring titin in the Z-disk is the actin cross-linker α-actinin. The calmodulin-like domain of α-actinin binds to the Z-repeats of titin. However, the mechanical and kinetic properties of this important interaction are still unknown. Here, we use a dual-beam optical tweezers assay to study the mechanics of this interaction at the single-molecule level. A single interaction of α-actinin and titin turns out to be surprisingly weak if force is applied. Depending on the direction of force application, the unbinding forces can more than triple. Our results suggest a model where multiple α-actinin/Z-repeat interactions cooperate to ensure long-term stable titin anchoring while allowing the individual components to exchange dynamically.

Nanomechanics of the substrate binding domain of Hsp70 determine its allosteric ATP-induced conformational change.

Owing to the cooperativity of protein structures, it is often almost impossible to identify independent subunits, flexible regions, or hinges simply by visual inspection of static snapshots. Here, we use single-molecule force experiments and simulations to apply tension across the substrate binding domain (SBD) of heat shock protein 70 (Hsp70) to pinpoint mechanical units and flexible hinges. The SBD consists of two nanomechanical units matching 3D structural parts, called the α- and ß-subdomain. We identified a flexible region within the rigid ß-subdomain that gives way under load, thus opening up the α/ß interface. In exactly this region, structural changes occur in the ATP-induced opening of Hsp70 to allow substrate exchange. Our results show that the SBD's ability to undergo large conformational changes is already encoded by passive mechanics of the individual elements.

An Active, Ligand-Responsive Pulling Geometry Reports on Internal Signaling between Subdomains of the DnaK Nucleotide-Binding Domain in Single-Molecule Mechanical Experiments.

Single-molecule mechanical experiments have proven to be ideal tools for probing the energetics and mechanics of large proteins and domains. In this paper, we investigate the nucleotide-dependent unfolding mechanics of the nucleotide-binding domain (NBD) of the Hsp70 chaperone DnaK. The NBD binds ADP or ATP in the binding cleft formed by lobe I and lobe II, which consists of two subdomains each. When force is applied to the termini of the NBD, the observed unfolding forces are independent of the nucleotide state. In contrast, when force is applied across the nucleotide-binding pocket, the unfolding forces report specifically on the nucleotide-phosphate state. In this active, ligand-responsive pulling geometry, we observed a bifurcation of the unfolding pathway; the pathway proceeds either through a cooperative "coupled pathway" or through a noncooperative "uncoupled pathway". The partitioning between individual unfolding pathways can be effectively tuned by mutation or by the nucleotide exchange factor GrpE, i.e., by the factors affecting the strength of the lobe I-lobe II interactions within the native NBD. These experiments provide important insight into the molecular origin of the internal signaling between the subdomains of the nucleotide-binding domain of Hsp70 proteins and how signals are efficiently transferred inside the protein molecule.

Patient Preferences for Coronary CT Angiography with Stress Perfusion, SPECT, or Invasive Coronary Angiography.

Background Patient preference is pivotal for widespread adoption of tests in clinical practice. Patient preferences for invasive versus other noninvasive tests for coronary artery disease are not known. Purpose To compare patient acceptance and preferences for noninvasive and invasive cardiac imaging in North and South America, Asia, and Europe. Materials and Methods This was a prospective 16-center trial in 381 study participants undergoing coronary CT angiography with stress perfusion, SPECT, and invasive coronary angiography (ICA). Patient preferences were collected by using a previously validated questionnaire translated into eight languages. Responses were converted to ordinal scales and were modeled with generalized linear mixed models. Results In patients in whom at least one test was associated with pain, CT and SPECT showed reduced median pain levels, reported on 0-100 visual analog scales, from 20 for ICA (interquartile range [IQR], 4-50) to 6 for CT (IQR, 0-27.5) and 5 for SPECT (IQR, 0-25) (P < .001). Patients from Asia reported significantly more pain than patients from other continents for ICA (median, 25; IQR, 10-50; P = .01), CT (median, 10; IQR, 0-30; P = .02), and SPECT (median, 7; IQR, 0-28; P = .03). Satisfaction with preparation differed by continent and test (P = .01), with patients from Asia reporting generally lower ratings. Patients from North America had greater percentages of "very high" or "high" satisfaction than patients from other continents for ICA (96% vs 82%, respectively; P < .001) and SPECT (95% vs 79%, respectively; P = .04) but not for CT (89% vs 86%, respectively; P = .70). Among all patients, CT was preferred by 54% of patients, compared with 18% for SPECT and 28% for ICA (P < .001). Conclusion For cardiac imaging, patients generally favored CT angiography with stress perfusion, while study participants from Asia generally reported lowest satisfaction. © RSNA, 2019 Online supplemental material is available for this article. See also the editorial by Woodard and Nguyen in this issue.

Noise reduction and motion elimination in low-dose 4D myocardial computed tomography perfusion (CTP): preliminary clinical evaluation of the ASTRA4D algorithm.

OBJECTIVES: To propose and evaluate a four-dimensional (4D) algorithm for joint motion elimination and spatiotemporal noise reduction in low-dose dynamic myocardial computed tomography perfusion (CTP). METHODS: Thirty patients with suspected or confirmed coronary artery disease were prospectively included and underwent dynamic contrast-enhanced 320-row CTP. A novel deformable image registration method based on the principal component analysis (PCA) of the ante hoc temporally smoothed voxel-wise time-attenuation curves (ASTRA4D) is presented. Quantitative (standard deviation, signal-to-noise ratio (SNR), temporal variation, volumetric deformation) and qualitative (motion, contrast, contour sharpness [1, poor; 5, excellent]) measures of CTP quality were assessed for the original and motion-compensated sequences (without and with temporal filtering, PCA/ASTRA4D). Following myocardial perfusion deficit detection by two readers, diagnostic accuracy was evaluated using magnetic resonance myocardial perfusion imaging (MR-MPI) as the reference standard in 15 patients. RESULTS: Registration using ASTRA4D was successful in all 30 patients and resulted in comparison with the benchmark PCA in significantly (p < 0.001) reduced noise over time (- 83%, 178.5 vs 29.9) and spatially (- 34%, 21.4 vs 14.1) as well as improved SNR (+ 47%, 3.6 vs 5.3) and subjective image quality (motion, contrast, contour sharpness [+ 1.0, + 1.0, + 0.5]). ASTRA4D had significantly improved per-segment sensitivity of 91% (58/64) and similar specificity of 96% (429/446) compared with PCA (52%, 33/64; 98%, 435/446; p = 0.011) in the visual detection of perfusion deficits. CONCLUSIONS: The ASTRA4D registration algorithm improved the spatiotemporal noise profile and CTP sequence image quality, resulting in significantly improved sensitivity of 4D CTP in the detection of myocardial ischemia. KEY POINTS: • ASTRA4D combines local temporal regression and deformable image registration. • Quantitative and qualitative measures of CTP quality are improved compared to PCA. • Improved spatiotemporal differentiation of ischemic regions leads to an excellent perfusion deficit concordance of ASTRA4D with MRI.

Folding and assembly of the large molecular machine Hsp90 studied in single-molecule experiments.

Folding of small proteins often occurs in a two-state manner and is well understood both experimentally and theoretically. However, many proteins are much larger and often populate misfolded states, complicating their folding process significantly. Here we study the complete folding and assembly process of the 1,418 amino acid, dimeric chaperone Hsp90 using single-molecule optical tweezers. Although the isolated C-terminal domain shows two-state folding, we find that the isolated N-terminal as well as the middle domain populate ensembles of fast-forming, misfolded states. These intradomain misfolds slow down folding by an order of magnitude. Modeling folding as a competition between productive and misfolding pathways allows us to fully describe the folding kinetics. Beyond intradomain misfolding, folding of the full-length protein is further slowed by the formation of interdomain misfolds, suggesting that with growing chain lengths, such misfolds will dominate folding kinetics. Interestingly, we find that small stretching forces applied to the chain can accelerate folding by preventing the formation of cross-domain misfolding intermediates by leading the protein along productive pathways to the native state. The same effect is achieved by cotranslational folding at the ribosome in vivo.

Knotting and unknotting of a protein in single molecule experiments.

Spontaneous folding of a polypeptide chain into a knotted structure remains one of the most puzzling and fascinating features of protein folding. The folding of knotted proteins is on the timescale of minutes and thus hard to reproduce with atomistic simulations that have been able to reproduce features of ultrafast folding in great detail. Furthermore, it is generally not possible to control the topology of the unfolded state. Single-molecule force spectroscopy is an ideal tool for overcoming this problem: by variation of pulling directions, we controlled the knotting topology of the unfolded state of the 52-knotted protein ubiquitin C-terminal hydrolase isoenzyme L1 (UCH-L1) and have therefore been able to quantify the influence of knotting on its folding rate. Here, we provide direct evidence that a threading event associated with formation of either a 31 or 52 knot, or a step closely associated with it, significantly slows down the folding of UCH-L1. The results of the optical tweezers experiments highlight the complex nature of the folding pathway, many additional intermediate structures being detected that cannot be resolved by intrinsic fluorescence. Mechanical stretching of knotted proteins is also of importance for understanding the possible implications of knots in proteins for cellular degradation. Compared with a simple 31 knot, we measure a significantly larger size for the 52 knot in the unfolded state that can be further tightened with higher forces. Our results highlight the potential difficulties in degrading a 52 knot compared with a 31 knot.

Coronary Artery Disease: Analysis of Diagnostic Performance of CT Perfusion and MR Perfusion Imaging in Comparison with Quantitative Coronary Angiography and SPECT-Multicenter Prospective Trial.

Purpose To compare the diagnostic performance of stress myocardial computed tomography (CT) perfusion with that of stress myocardial magnetic resonance (MR) perfusion imaging in the detection of coronary artery disease (CAD). Materials and Methods All patients gave written informed consent prior to inclusion in this institutional review board-approved study. This two-center substudy of the prospective Combined Noninvasive Coronary Angiography and Myocardial Perfusion Imaging Using 320-Detector Row Computed Tomography (CORE320) multicenter trial included 92 patients (mean age, 63.1 years ± 8.1 [standard deviation]; 73% male). All patients underwent perfusion CT and perfusion MR imaging with either adenosine or regadenoson stress. The predefined reference standards were combined quantitative coronary angiography (QCA) and single-photon emission CT (SPECT) or QCA alone. Results from coronary CT angiography were not included, and diagnostic performance was evaluated with the Mantel-Haenszel test stratified by disease status. Results The prevalence of CAD was 39% (36 of 92) according to QCA and SPECT and 64% (59 of 92) according to QCA alone. When compared with QCA and SPECT, per-patient diagnostic accuracy of perfusion CT and perfusion MR imaging was 63% (58 of 92) and 75% (69 of 92), respectively (P = .11); sensitivity was 92% (33 of 36) and 83% (30 of 36), respectively (P = .45); and specificity was 45% (25 of 56) and 70% (39 of 56), respectively (P < .01). When compared with QCA alone, diagnostic accuracy of CT perfusion and MR perfusion imaging was 82% (75 of 92) and 74% (68 of 92), respectively (P = .27); sensitivity was 90% (53 of 59) and 69% (41 of 59), respectively (P < .01); and specificity was 67% (22 of 33) and 82% (27 of 33), respectively (P = .27). Conclusion This multicenter study shows that the diagnostic performance of perfusion CT is similar to that of perfusion MR imaging in the detection of CAD. © RSNA, 2017 Online supplemental material is available for this article.