MQ-HNCO-TROSY for the measurement of scalar and residual dipolar couplings in larger proteins: application to a 557-residue IgFLNa16-21

We describe a novel pulse sequence, MQ-HNCO-TROSY, for the measurement of scalar and residual dipolar couplings between amide proton and nitrogen in larger proteins. The experiment utilizes the whole 2T_N polarization transfer delay for labeling of ¹⁵N chemical shift in a constant time manner, which efficiently doubles the attainable resolution in ¹⁵N dimension with respect to the conventional HNCO-TROSY experiment. In addition, the accordion principle is employed for measuring (J + D)_NHs, and the multiplet components are selected with the generalized version of the TROSY scheme introduced by Nietlispach (J Biomol NMR 31:161–166, 2005). Therefore, cross peak overlap is diminished while the time period during which the ¹⁵N spin is susceptible to fast transverse relaxation associated with the anti-TROSY transition is minimized per attainable resolution unit. The proposed MQ-HNCO-TROSY scheme was employed for measuring RDCs in high molecular weight protein IgFLNa16-21 of 557 residues, resulting in 431 experimental RDCs. Correlations between experimental and back-calculated RDCs in individual domains gave relatively low Q-factors (0.19–0.39), indicative of sufficient accuracy that can be obtained with the proposed MQ-HNCO-TROSY experiment in high molecular weight proteins.     

Suppression of anti-TROSY lines in a sensitivity enhanced gradient selection TROSY scheme

A simple modification of the TROSY pulse transfer scheme, suggested by Yang and Kay [J. Biomol. NMR 13 (1999) 3–10], is proposed which results in the suppression of unwanted anti-TROSY lines without any extra loss in sensitivity. The higher sensitivity of this TROSY transfer scheme therefore becomes available for 2D [¹⁵N, ¹H] TROSY correlation and 3D/4D ¹⁵N separated NOESY type experiments where complete suppression of the broad anti-TROSY lines is essential.     

TROSY pulse sequence for simultaneous measurement of the <Superscript>15</Superscript>N <Emphasis Type="Italic">R</Emphasis><Subscript>1</Subscript> and {<Superscript>1</Superscript>H}–<Superscript>15</Superscript>N NOE in deuterated proteins

A TROSY-based NMR experiment is described for simultaneous measurement of the ¹⁵N longitudinal relaxation rate constant R₁ and the {¹H}–¹⁵N nuclear Overhauser enhancement. The experiment is based on the observation that the TROSY mixing pulse sequence element symmetrically exchanges ¹H and ¹⁵N magnetizations. The accuracy of the proposed technique is validated by comparison to independent measurements of both relaxation parameters for the protein ubiquitin. The simultaneous experiment is approximately 20–33% shorter than conventional sequential measurements.     

Tuning the HNN experiment: generation of serine–threonine check points

We describe here the tunability of the HNN experiment to obtain certain residue specific peak patterns in the spectra of (¹⁵N, ¹³C) labeled proteins. This is achieved by tuning a band-selective 180° pulse on the carbon channel in the pulse sequence, whereby one can tamper with the C^α–C^β coupling evolutions for the different residues. Specifically, we generate distinctive peak patterns for serine and threonine and their neighbors in the different planes of the three dimensional spectrum. These provide useful anchor points during sequential assignment of backbone resonances. The performance of this experiment, referred to as HNN-ST here, is demonstrated using two proteins, one properly folded and the other completely denatured. With the availability of high field spectrometers, techniques such as TROSY, and ever increasing sensitivities in the probes, this experiment with its large number of check points has a great potential for rapid and unambiguous backbone resonance assignment in large proteins.     

A TROSY CPMG sequence for characterizing chemical exchange in large proteins

A new NMR spin relaxation experiment is described for measuring chemical exchange time constants from approximately 0.5 ms to 5 ms in ¹⁵N-labeled macromolecules. The pulse sequence is based on the Carr–Purcell–Meiboom–Gill technique [Carr and Purcell (1954) Phys. Rev., 94, 630–638; Meiboom and Gill (1958) Rev. Sci. Instrum., 29, 688–691; Loria et al. (1999) J. Am. Chem. Soc., 121, 2331–2332], but implements TROSY selection [Pervushin et al. (1997) Proc. Natl. Acad. Sci. USA, 94, 12366–12371] to permit measurement of exchange linebroadening contributions to the narrower component of the ¹H-¹⁵N scalar-coupled doublet. This modification extends the size limitation imposed on relaxation measurements due to the fast decay of transverse magnetization in larger macromolecules. The new TROSY-CPMG experiment is demonstrated on a [U-98% ¹⁵ N] labeled sample of basic pancreatic trypsin inhibitor and a [U-83% ²H, U-98% ¹⁵ N] labeled sample of triosephosphate isomerase, a 54 kDa homodimeric protein.     

Molecular cloning,expression, and cytokinin (6-benzylaminopurine) antagonist activity of peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis>) lectin SL-I

Isolation and purification of a α-methyl-mannoside specific lectin (SL-I) of peanut was reported earlier [Singh and Das (1994) Glycoconj J 11:282–285]. Native SL-I is a glycoprotein having ∼31 kDa subunit molecular mass and forms dimer. The gene encoding this lectin is identified from a 6-day old peanut root cDNA library by anti-SL-I antibody and N-terminal amino acid sequence homology to the native lectin. Nucleotide sequence derived amino acid sequence of the re-SL-I shows amino acid sequence homology with the N-terminal and tryptic digests’ amino acid sequence of the native SL-I (nSL-I). Presence of a putative glycosylation (QNPS) site and a hydrophobic adenine-binding (VLVSYDANS) site is also identified in SL-I. Homology modeling of the lectin suggests it to be an archetype of legume lectins. It is expressed as a ~30 kDa apoprotein in E. coli and has the carbohydrate specificity and secondary structure identical to its natural counterpart. The lectin SL-I inhibits cytokinin 6-benzylaminopurine (BA)-induced “delayed leaf senescence” and “cotyledon expansion”. Equilibrium dialysis revealed a single high-affinity binding site for adenine (7.6 × 10⁻⁶ M) and BA (1.09 × 10⁻⁵ M) in the SL-I dimer and thus suggesting that the cytokinin antagonist effect of SL-I is mediated by the direct interaction of SL-I with BA.The nucleotide sequence data reported here are available in the DDBJ/EMBL/GenBank databases under the Accession No. AJ585523     

Perspective: revisiting the field dependence of TROSY sensitivity

The discovery of the TROSY effect (Pervushin et al. in Proc Natl Acad Sci USA 94:12366–12371, 1997) for reducing transverse relaxation and line sharpening through selecting pathways in which dipole–dipole and CSA Hamiltonians partially cancel each other had a tremendous impact on solution NMR studies of macromolecules. Together with the methyl TROSY (Tugarinov and Kay in J Biomol NMR 28:165–172, 2004) it enabled structural and functional studies of significantly larger systems. The optimal field strengths for TROSY have been estimated to be on spectrometers operating around 900 MHz (21.14 T) for the ¹H_N TROSY (Pervushin et al. in Proc Natl Acad Sci USA 94:12366–12371, 1997) while the aromatic ¹³C (¹³C_aro) TROSY is posited to be optimal at around 600 MHz (14.09 T) (Pervushin et al. in J Am Chem Soc 120:6394–6400, 1998b; Pervushin in Q Rev Biophys 33:161–197, 2000). The initial rational was based on the consideration of where the quadratic B₀ field dependences of the TROSY relaxation rates reach a minimum. For sensitivity consideration, however, it is interesting to estimate which field strengths yield the tallest peaks. Recent studies of ¹⁵N-detected TROSYs suggested that maximal peak heights are expected at 1.15 GHz (27.01 T) although the slowest relaxation rates or longest transverse relaxation times T₂ are indeed expected around 900 MHz (21.14 T) (Takeuchi in J Biomol NMR 63:323–331, 2015; Takeuchi et al. in J Biomol NMR 64:143–151, 2016). This was based on the fact that the heights of Lorentzian lines are proportional to B _o ^3/2 * T₂ (B_o). Thus, multiplying the parabolic T₂(B_o) dependence with the increasing function of B _o ^3/2 shifts the maxima of peak-height field dependence from the T₂ maximum at 900 MHz to higher fields. Moreover, besides shifting the peak height maximum for ¹⁵N TROSY, this analysis yields estimates for optimal peak heights for ¹H_N detected TROSY to 1.5 GHz, and to 900 MHz for ¹³C-detected ¹³C_aroTROSY as is detailed below. To our knowledge, this aspect of field dependence of TROSY sensitivity has not been in the attention of the NMR community but may affect perspectives of NMR at ultra-high fields.     

Speeding up direct <Superscript>15</Superscript>N detection: hCaN 2D NMR experiment

Experiments detecting low gyromagnetic nuclei have recently been proposed to utilize the relatively slow relaxation properties of these nuclei in comparison to ¹H. Here we present a new type of ¹⁵N direct-detection experiment. Like the previously proposed CaN experiment (Takeuchi et al. in J Biomol NMR 47:271–282, 2010), the hCaN experiment described here sequentially connects amide ¹⁵N resonances, but utilizes the initial high polarization and the faster recovery of the ¹H nucleus to shorten the recycling delay. This allows recording 2D ¹⁵N-detected NMR experiments on proteins within a few hours, while still obtaining superior resolution for ¹³C and ¹⁵N, establishing sequential assignments through prolines, and at conditions where amide protons exchange rapidly. The experiments are demonstrated on various biomolecules, including the small globular protein GB1, the 22 kDa HEAT2 domain of eIF4G, and an unstructured polypeptide fragment of NFAT1, which contains many SerPro sequence repeats.     

Ebbie: automated analysis and storage of small RNA cloning data using a dynamic web server

Background  

DNA sequencing is used ubiquitously: from deciphering genomes[1] to determining the primary sequence of small RNAs (smRNAs) [2–5]. The cloning of smRNAs is currently the most conventional method to determine the actual sequence of these important regulators of gene expression. Typical smRNA cloning projects involve the sequencing of hundreds to thousands of smRNA clones that are delimited at their 5' and 3' ends by fixed sequence regions. These primers result from the biochemical protocol used to isolate and convert the smRNA into clonable PCR products. Recently we completed a smRNA cloning project involving tobacco plants, where analysis was required for ~700 smRNA sequences[6]. Finding no easily accessible research tool to enter and analyze smRNA sequences we developed Ebbie to assist us with our study.     

Intermediate binding of phycocyanobilin to the lyase, CpeS1, and transfer to apoprotein

The phycobilin: Cysteine-84-phycobiliprotein lyase, CpeS1, catalyzes phycocyanobilin (PCB) and phycoerythrobilin attachment to nearly all cysteine-84 (consensus sequence) binding sites of phycoerythrin, phycoerythrocyanin, phycocyanin and allophycocyanin (Zhao et al. (2007) Proc Natl Acad Sci 104:14300–14305). We now show that CpeS1 can bind PCB, as assayed by Ni²⁺ chelating affinity chromatography. Binding is rapid, and the chromophore is bound in an extended conformation similar to that in phycobiliproteins but only poorly fluorescent. Upon addition of apo-biliproteins, the chromophore is transferred to the latter much slower (∼1 h), indicating that chromophorylated CpeS1 is an intermediate in the enzymatic reaction. In addition, imidazole is bound to PCB, as shown by mass spectroscopy of tryptic digests of the intermediate CpeS1–PCB complex.     

Induction of a non-specific permeability transition in mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts

In this study we used tightly-coupled mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts, possessing a respiratory chain with the usual three points of energy conservation. High-amplitude swelling and collapse of the membrane potential were used as parameters for demonstrating induction of the mitochondrial permeability transition due to opening of a pore (mPTP). Mitochondria from Y. lipolytica, lacking a natural mitochondrial Ca²⁺ uptake pathway, and from D. magnusii, harboring a high-capacitive, regulated mitochondrial Ca²⁺ transport system (Bazhenova et al. J Biol Chem 273:4372–4377, 1998a; Bazhenova et al. Biochim Biophys Acta 1371:96–100, 1998b; Deryabina and Zvyagilskaya Biochemistry (Moscow) 65:1352–1356, 2000; Deryabina et al. J Biol Chem 276:47801–47806, 2001) were very resistant to Ca²⁺ overload. However, exposure of yeast mitochondria to 50–100 μM Ca²⁺ in the presence of the Ca²⁺ ionophore ETH129 induced collapse of the membrane potential, possibly due to activation of the fatty acid-dependent Ca²⁺/nH⁺-antiporter, with no classical mPTP induction. The absence of response in yeast mitochondria was not simply due to structural limitations, since large-amplitude swelling occurred in the presence of alamethicin, a hydrophobic, helical peptide, forming voltage-sensitive ion channels in lipid membranes. Ca²⁺- ETH129-induced activation of the Ca²⁺/H⁺-antiport system was inhibited and prevented by bovine serum albumin, and partially by inorganic phosphate and ATP. We subjected yeast mitochondria to other conditions known to induce the permeability transition in animal mitochondria, i.e., Ca²⁺ overload (in the presence of ETH129) combined with palmitic acid (Mironova et al. J Bioenerg Biomembr 33:319–331, 2001; Sultan and Sokolove Arch Biochem Biophys 386:37–51, 2001), SH-reagents, carboxyatractyloside (an inhibitor of the ADP/ATP translocator), depletion of intramitochondrial adenine nucleotide pools, deenergization of mitochondria, and shifting to acidic pH values in the presence of high phosphate concentrations. None of the above-mentioned substances or conditions induced a mPTP-like pore. It is thus evident that the permeability transition in yeast mitochondria is not coupled with Ca²⁺ uptake and is differently regulated compared to the mPTP of animal mitochondria.     

The taxonomy and phylogeny of the cyprinid genus Opsariichthys Bleeker (Teleostei: Cyprinidae) from Taiwan, with description of a new species

The morphological and mitochondrial genetic differentiation in the cyprinid genus, Opsariichthys Bleeker (Nederlandsch Tijdschrift voor de Dierkunde 1:187–218, 1863) have been surveyed in Taiwan. Among them, there are three valid species can be recognized in Taiwan including Opsariichthys pachycephalus Günther (1868) distributed in northern and western Taiwan, Opsariichthys evolans (Jordan and Evermann Proc US Nat Mus 25:315–368, 1902) in northern Taiwan and an unnamed species from southern Taiwan described herein as Opsariichthys kaopingensis Chen and Wu, new species which can be well distinguished from the related O. pachycaphalus by their body proportions, scale counts, and specific coloration patterns. We utilized mitochondrial complete D-loop sequence data to infer phylogenetic relationships within a subset of related genera of opsariichthines, and to examine evidence for genetic differentiation in these two sibling species formerly assigned to “Zacco” pachycephalus and their genetic relationship with other congeneric species around nearby regions. The clade of O. pachycephalus and O. kaopingensis in genetically were recovered as more closely related to Opsariichthys uncirostris (Temminck and Schlegel 1846) species complex including both O. uncirostris and O. bidens Günther (1868) from Japan and mainland China than to typical Zacco from Japan. This molecular phylogenetic insight strongly supports the assignment for both so-called “Zacco” pachycephalus and this new species described herein as the typical monophyletic members of Opsariichthys and the type species of Zacco as Zacco platypus (Temminck and Schlegel 1846) from Japan is sister clade for all species groups in Opsariichthys. Opsariichthys pachycephalus and O. kaopingensis were strongly differentiated by large mitogenetic distances and phylogenetic support from distance and discrete method and Bayesian inference based on complete mtDNA D-loop sequences, with an average mitogenetic divergence of 3.3%, which may suggest that the separation of the two species happened much earlier than the last glacial period. Opsariichthys evolans seems to share the close genetic relationship with O. acutipinnis (Bleeker Nederlandsch Tijdschrift voor de Dierkunde 1:187–218, 1863) from the Yangtsi River basin.     

Hadamard Phylogenetic Methods and the <Emphasis Type="Italic">n</Emphasis>-taxon Process

The Hadamard transform (Hendy and Penny, Syst. Zool. 38(4):297–309, 1989; Hendy, Syst. Zool. 38(4):310–321, 1989) provides a way to work with stochastic models for sequence evolution without having to deal with the complications of tree space and the graphical structure of trees. Here we demonstrate that the transform can be expressed in terms of the familiar P[τ]=e ^Q[τ] formula for Markov chains. The key idea is to study the evolution of vectors of states, one vector entry for each taxa; we call this the n-taxon process. We derive transition probabilities for the process. Significantly, the findings show that tree-based models are indeed in the family of (multi-variate) exponential distributions.     

Evidence for Na+–glucose cotransporter in type I alveolar epithelium

Functional evidence of Na⁺–glucose cotransport in rat lung has been provided by Basset et al. (J. Physiol. 384:325–345, 1987). By autoradiography [³H]phloridzin binding has been found confined to alveolar epithelial type II cells in mouse and rabbit lungs (Boyd, J. Physiol. 422: 44P, 1990). In this research we checked by immunofluorescence whether Na⁺–glucose cotransporter (SGLT1) is also expressed in alveolar type I cells. Lungs of anesthetized rats and lambs were fixed by paraformaldehyde, perfused in pulmonary artery, or instilled into a bronchus, respectively. Tissue blocks embedded in paraffin or frozen were sectioned. Two specific anti-SGLT1 antibodies for rat recognizing aminoacid sequence 402–420, and 546–596 were used in both species. Bound primary antibody was detected by secondary antibody conjugated to fluorescein isothiocianate or Texas red, respectively. In some sections cellular nuclei were also stained. In rats alveolar type I cells were identified by fluorescent Erythrina cristagalli lectin. Sections were examined by confocal laser-scanning microscope. Both in rats and lambs alveolar epithelium was stained by either antibody; no labeling occurred in negative controls. Hence, SGLT1 appears to be also expressed in alveolar type I cells. This is functionally relevant because type I cells provide 95–97% of alveolar surface, and SGLT1, besides contributing to removal of lung liquid under some circumstances, keeps low glucose concentration in lining liquid, which is useful to prevent lung infection.     

qTROSY – a novel scheme for recovery of the anti-TROSY magnetisation

In TROSY experiments, spin state selection (S³) retains only the single HSQC sub-spectrum with minimal T₂ relaxation and maximal resolution, yet at the cost of eliminating half of the available polarisation as undesired anti-TROSY component. We here introduce queued TROSY (qTROSY) as a novel scheme to partially recover and exploit this anti-TROSY polarisation in two concatenated scans. After initial orthogonal spin state separation (oS³), anti-TROSY polarisation is explicitly stored while its TROSY counterpart follows the desired coherence pathway recorded in a first scan A. The immediately appended scan B then quantitatively converts the recovered anti-TROSY polarisation into a second TROSY spectrum, skipping the time-limiting long reequilibration delay. Both concatenated qTROSY scans thus ideally exploit the full initial polarisation within almost the same measurement time. In practice, T₂ relaxation losses accruing during the coupling evolution delays reduced anti-TROSY polarisation recovery below 40%, obviating sensitivity enhancement through addition of both qTROSY scans; yet, scan B retained a complete scan A spectrum with up to 75% intensity. We therefore propose to employ qTROSY asymmetrically, compacting two separate conventional into one queued TROSY-type experiment with significantly reduced measurement time, implying primarily the concatenation of different three- or higher-dimensional experiments. Both anti-TROSY polarisation recoveries and possible time savings are largest for deuterated and smaller non-deuterated proteins, extending the rentability limit of the TROSY principle towards smaller molecular weights.Supplementary material to this paper is available in electronic form at http://dx.doi.org/10.1007/s10858-005-5618-z     

100% complete assignment of non-labile 1H, 13C, and 15N signals for calcium-loaded calbindin D9k P43G

Here we present the 100% complete assignment chemical shift of non-labile ¹H, ¹⁵N and ¹³C nuclei of Calbindin D_9k P43G. The assignment includes all non-exchangeable side chain nuclei, including ones that are rarely reported, such as LysNζ as well as the termini. NMR experiments required to achieve truly complete assignments are discussed. To the best of our knowledge our assignments for Calbindin D_9k extend beyond previous studies reaching near-completeness (Vis et al. in Biochem 33:14858–14870, 1994; Yamazaki et al. in J Am Chem Soc 116:6464–6465, 1994; Yamazaki et al. in Biochem 32:5656–5669, 1993b).     

Accuracy of dominant markers for estimation of relatedness and heritability in an experimental stand of Prosopis alba (Leguminosae)

The estimation of genetic components of phenotypic variance is based on the resemblance between relatives. In natural populations of most forest tree species without genealogical information, a possible alternative approach is the use of relatedness estimates obtained indirectly from molecular marker data. Heritability (h ²) is then estimated from the covariance of estimated relatedness and phenotypic resemblance. In a stand of Prosopis alba planted in 1991 in Argentina, relatedness was estimated for all individual pairs of trees by means of the information proceeding from 128 dominant markers (57 AFLPs and 71 ISSRs) and compared with estimates obtained from six microsatellite loci previously studied. We empirically compared the accuracy of different relatedness estimators based on dominant markers proposed by Lynch and Milligan (Mol Ecol 3:91–99, 1994), Hardy (Mol Ecol 12:1577–1588, 2003), Wang (Mol Ecol 13:3169–3178, 2004), and Ritland (Mol Ecol 14:3157–3165, 2005). Heritabilities of 13 quantitative traits were then estimated from the regression of pairwise phenotypic distances on pairwise relatedness according to Ritland (Genet Res 67:175–185, 1996a). Relatedness inferred from molecular markers was in all cases significantly correlated with actual relatedness, although Ritland's estimator showed the highest bias but the lowest variance. Dominant marker-based h ² estimates were evidently downwards biased and showed poor correlation with those based on family records. In conclusion, the use of dominant molecular markers evidently produces much greater underestimates of h ² than from using co-dominant ones, attributable to the lower accuracy in the indirect estimation of relatedness coefficient. Many traits with enough genetic variability as to respond readily to selection would remain undetected; only those with very high heritability would show significant h ² estimates.     

HNHC: a triple resonance experiment for correlating the H2, N1(N3) and C2 resonances in adenine nucleobases of <Superscript>13</Superscript>C-, <Superscript>15</Superscript>N-labeled RNA oligonucleotides

A novel NMR pulse sequence has been developed that correlates the H2 resonances with the C2 and the N1 (N3) resonances in adenine nucleobases of ¹³C, ¹⁵N labeled oligonucleotides. The pulse scheme of the new 3D-HNHC experiment is composed of a ²J-¹⁵N-HSQC and a ¹J-¹³C-HSQC and utilizes large ²J(H2, N1(N3)) and ¹J(H2, C2) couplings. The experiment was applied to a medium-size ¹³C, ¹⁵N-labeled 36mer RNA. It is useful to resolve assignment ambiguities occurring especially in larger RNA molecules due to resonance overlap in the ¹H-dimension. Therefore, the missing link in correlating the imino H3 resonances of the uracils across the AU base pair to the H8 resonances of the adenines via the novel pulse sequence and the TROSY relayed HCCH-COSY (Simon et al. in J Biomol NMR 20:173–176 2001) is provided. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Kinetic models of photosystem II should accommodate the effect of donor side quenching on variable chlorophyll <Emphasis Type="Italic">a</Emphasis> fluorescence in the microseconds time range

Quantitative data on laser flash-induced variable fluorescence in the 100 ns to 1 ms time range (Belyaeva et al. in Photosynth Res 98:105–119, 2008) confirming those of others (Steffen et al. in Biochemistry 40:173–180, 2001, Biochemistry 44:3123–3132, 2005; Belyaeva et al. in Biophysics 51(6):976–990, 2006), need a substantial correction with respect to magnitude of the normalized variable fluorescence associated with single turnover-induced charge separation in RCs of PS II. Their data are conclusive with the involvement of donor side quenching, the release of which occurs with a rate constant in the range of tens of ms⁻¹, and presumed to be associated with reduction of Y_\textz ⁺ Y_{\text{z}}^{ + } by the OEC.     

1H, 13C and 15N NMR assignments of the C1A and C1B subdomains of PKC-delta

The Protein Kinase C family of enzymes is a group of serine/threonine kinases that play central roles in cell-cycle regulation, development and cancer. A key step in the activation of PKC is translocation to membranes and binding of membrane-associated activators including diacylglycerol (DAG). Interaction of novel and conventional isotypes of PKC with DAG and phorbol esters occurs through the two C1 regulatory domains (C1A and C1B), which exhibit distinct ligand binding selectivity that likely controls enzyme activation by different co-activators. PKC has also been implicated in physiological responses to alcohol consumption and it has been proposed that PKCα (Slater et al. J Biol Chem 272(10):6167–6173, 1997; Slater et al. Biochemistry 43(23):7601–7609, 2004), PKCε (Das et al. Biochem J 421(3):405–413, 2009) and PKCδ (Das et al. J Biol Chem 279(36):37964–37972, 2004; Das et al. Protein Sci 15(9):2107–2119, 2006) contain specific alcohol-binding sites in their C1 domains. We are interested in understanding how ethanol affects signal transduction processes through its affects on the structure and function of the C1 domains of PKC. Here we present the ¹H, ¹⁵N and ¹³C NMR chemical shift assignments for the Rattus norvegicus PKCδ C1A and C1B proteins.