The register shift rules for βαβ-motifs for de novo protein design

A wide range of de novo design of αβ-proteins has been achieved based on the design rules, which describe secondary structure lengths and loop torsion patterns favorable for design target topologies. This paper proposes design rules for register shifts in βαβ-motifs, which have not been reported previously, but are necessary for determining a target structure of de novo design of αβ-proteins. By analyzing naturally occurring protein structures in a database, we found preferences for register shifts in βαβ-motifs, and derived the following empirical rules: (1) register shifts must not be negative regardless of torsion types for a constituent loop in βαβ-motifs; (2) preferred register shifts strongly depend on the loop torsion types. To explain these empirical rules by physical interactions, we conducted physics-based simulations for systems mimicking a βαβ-motif that contains the most frequently observed loop type in the database. We performed an exhaustive conformational sampling of the loop region, imposing the exclusion volume and hydrogen bond satisfaction condition. The distributions of register shifts obtained from the simulations agreed well with those of the database analysis, indicating that the empirical rules are a consequence of physical interactions, rather than an evolutionary sampling bias. Our proposed design rules will serve as a guide to making appropriate target structures for the de novo design of αβ-proteins.

“Register-shift” insulin analogs uncover constraints of proteotoxicity in protein evolution

Globular protein sequences encode not only functional structures (the native state) but also protein foldability, i.e. a conformational search that is both efficient and robustly minimizes misfolding. Studies of mutations associated with toxic misfolding have yielded insights into molecular determinants of protein foldability. Of particular interest are residues that are conserved yet dispensable in the native state. Here, we exploited the mutant proinsulin syndrome (a major cause of permanent neonatal-onset diabetes mellitus) to investigate whether toxic misfolding poses an evolutionary constraint. Our experiments focused on an invariant aromatic motif (PheB24–PheB25–TyrB26) with complementary roles in native self-assembly and receptor binding. A novel class of mutations provided evidence that insulin can bind to the insulin receptor (IR) in two different modes, distinguished by a “register shift” in this motif, as visualized by molecular dynamics (MD) simulations. Register-shift variants are active but defective in cellular foldability and exquisitely susceptible to fibrillation in vitro. Indeed, expression of the corresponding proinsulin variant induced endoplasmic reticulum stress, a general feature of the mutant proinsulin syndrome. Although not present among vertebrate insulin and insulin-like sequences, a prototypical variant ([GlyB24]insulin) was as potent as WT insulin in a rat model of diabetes. Although in MD simulations the shifted register of receptor engagement is compatible with the structure and allosteric reorganization of the IR-signaling complex, our results suggest that this binding mode is associated with toxic misfolding and so is disallowed in evolution. The implicit threat of proteotoxicity limits sequence variation among vertebrate insulins and insulin-like growth factors.

Perceptual cues for individual voice quality

The alteration of phonation modes may affect speaker identification, but it is not known to what extent changes in register can compromise discrimination in each case. To answer this question, specifically with respect to falsetto voice, a perception experiment was designed using disguised voice in order to analyze the listeners’ reaction to stimuli that differed only with respect to phonation modes. Results were significant, suggesting that falsetto voice does not compromise speaker recognition. In addition to the perception experiment, the chapter also provides a detailed analysis of the temporal aspects of each mode and the biomechanical behavior of the vocal folds. Connecting the perception experiment to the production analysis allows us, first, to know which parameters have been altered, and which have not, during the register shift to falsetto voice and, secondly, to discuss what perceptive weight such parameters may have on the recognition of individual voice quality.

Phonetic realisation of downstep in Bimoba

Few phonological phenomena have so captured the attention of theorists and continued to baffle them as the phenomenon of tonal downstep. Downstep is the lowering of the tonal register that sometimes occurs between adjacent, otherwise identical tones. It is cumulative, and successive occurrences of the phenomenon result in ever lower settings of the tonal register. The present work reports on an instrumental study of downstep in Bimoba, a Gur language spoken in the Northern Region of Ghana. So far as I am aware, the present work is the only description in existence of tone in Bimoba.Bimoba is a good candidate for a study of downstep, because it is a ‘three-tone’ language in which both Low tones (floating and non-floating) and Mid tones cause High tones to be downstepped. A number of phonological studies (based on auditory impressions only) of three-tone languages with downstep of High tone have claimed that the difference in pitch between a High and a following Mid is equivalent to that between a High and a following downstepped High. These include Supyire (Carlson 1983), Babanki (Hyman 1979), Moba (Russell 1986) and Kagoro (van de Kolk 1992). This issue is important to tone theorists because a number of proposals on the theory of downstep formally equate a downstepped High with one type of Mid tone. These include Clements (1983), Hyman (1986, 1993), Inkelas (1987) and Snider (1990). On the other hand, a number of other proposals account for downstep with phonetic implementation rules. These include Pierrehumbert (1980), Poser (1984), Beckman & Pierrehumbert (1986) and Pierrehumbert & Beckman (1988). ‘If register shift is indeed a rule of phonetic implementation, there is no reason, a priori, for the interval of register shift to be equivalent to that found between two phonemic tones’ (Snider 1990: 470). So far as I am aware, no instrumental study has ever addressed this issue. The present study therefore undertakes to help fill this lacuna, and concludes that in Bimoba a Mid tone is phonetically indistinguishable from a downstepped High tone in comparable environments.