An exceptionally stable helix from the ribosomal protein L9: implications for protein folding and stability

Kuhlman, Brian; Yang, Hui; Boice, Judith A.; Fairman, Robert; Raleigh, Daniel P.

doi:10.1006/jmbi.1997.1146

Cited by 49 publications

(48 citation statements)

References 51 publications

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“…We also found that the thermal and salt stability of this peptide is similar to that described for other single stable ␣-helices (17)(18)(19)(20). Heating to 80°C reduced the helical content in a non-cooperative manner (Fig.…”

Section: Resultssupporting

confidence: 55%

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The Predicted Coiled-coil Domain of Myosin 10 Forms a Novel Elongated Domain That Lengthens the Head

Knight

Thirumurugan

et al. 2005

Journal of Biological Chemistry

140

186

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Myosin 10 contains a region of predicted coiled coil 120 residues long. However, the highly charged nature and pattern of charges in the proximal 36 residues appear incompatible with coiled-coil formation. Circular dichroism, NMR, and analytical ultracentrifugation show that a synthesized peptide containing this region forms a stable single ␣-helix (SAH) domain in solution and does not dimerize to form a coiled coil even at millimolar concentrations. Additionally, electron microscopy of a recombinant myosin 10 containing the motor, the three calmodulin binding domains, and the fulllength predicted coiled coil showed that it was mostly monomeric at physiological protein concentration. In dimers the molecules were joined only at their extreme distal ends, and no coiled-coil tail was visible. Furthermore, the neck lengths of both monomers and dimers were much longer than expected from the number of calmodulin binding domains. In contrast, micrographs of myosin 5 heavy meromyosin obtained under the same conditions clearly showed a coiled-coil tail, and the necks were the predicted length. Thus the predicted coiled coil of myosin 10 forms a novel elongated structure in which the proximal region is a SAH domain and the distal region is a SAH domain (or has an unknown extended structure) that dimerizes only at its end. Sequence comparisons show that similar structures may exist in the predicted coiled-coil domains of myosins 6 and 7a and MyoM and could function to increase the size of the working stroke.Myosins make up a diverse superfamily of motor proteins (1). The human genome alone contains about 40 myosin genes (2). Of these, about one-third are "conventional" myosins (i.e. the well studied myosin 2), and the rest fall into about 10 different classes. The structure, properties and functions of the majority of myosin classes are poorly characterized and have largely been inferred from sequence comparisons rather than direct experiments on purified proteins (1-3).Muscle myosin 2 dimerizes through its ␣-helical coiled-coil tail. Therefore, it is commonly assumed that any myosin will also be dimeric if it contains a region predicted to be coiled coil. This assumption is dependent on the accuracy of coiled-coil prediction programs, such as COILS (4), PAIRCOIL, or MULTICOIL (5), which are also used by protein-fold prediction sites on the Web such as SMART (6).Although myosin 10 contains a region of predicted coiled coil (Fig. 1A), and is predicted to dimerize, this has not been determined experimentally. We noticed that part of the predicted coiled-coil domain of myosin 10 is highly enriched in charged residues (Fig. 1B). The proximal region consisting of 36 residues is particularly enriched in both positively and negatively charged residues, including the a and d positions of the heptad repeat (a-g) that are canonically hydrophobic residues in coiled coils (Fig. 1B). We suspected that this highly charged sequence is unlikely to form a coiled coil (7), suggesting that this part of myosin 10 may not be able to dimerize....

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Section: Resultssupporting

confidence: 55%

“…1, C (solid lines) and D). SAH domains are rare but have been described for a small number of proteins including smooth muscle caldesmon (17) and the ribosomal protein L9 (18). The heptad repeat in the predicted SAH domain in myosin 10 has the novel general form ((R/K)XXXEXX) n .…”

Section: Resultsmentioning

confidence: 99%

The Predicted Coiled-coil Domain of Myosin 10 Forms a Novel Elongated Domain That Lengthens the Head

Knight

Thirumurugan

et al. 2005

Journal of Biological Chemistry

140

186

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“…The structure of the protein has been shown to be preserved in solution (Lillemoen et al, 1997). Studies of a peptide consisting of the long central helix show that it is monomeric and essentially fully structured in isolation, thus providing additional evidence that the unusual structure of L9 is preserved in solution (Kuhlman et al, 1997).…”

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confidence: 98%

Amide proton exchange measurements as a probe of the stability and dynamics of the n‐terminal domain of the ribosomal protein L9: Comparison with the intact protein

1998

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Amide H/D exchange rates have been measured for the N-terminal domain of the ribosomal protein L9, residues 1-56. The rates were measured at pD 3.91, 5.03, and 5.37. At pD 5.37, 18 amides exchange slowly enough to give reliable rate measurements. At pD 3.91, seven additional residues could be followed. The exchange is shown to occur by the EX2 mechanism for all conditions studied. The rates for the N-terminal domain are very similar to those previously measured for the corresponding region in the full-length protein (Lillemoen J et al., 1997, J Mol Biol 268:482-493). In particular, the rates for the residues that we have shown to exchange via global unfolding in the N-terminal domain agree within the experimental error with the rates measured by Hoffman and coworkers, suggesting that the structure of the domain is preserved in isolation and that the stability of the isolated domain is comparable to the stability of this domain in intact L9.

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“…In contrast, SAH domains are characterized by sequences that are rich in E, K and R residues and intra-molecular interactions between E and K and E and R in successive turns of the helix stabilise the helix in isolation [16]. SAH domains were first described some time ago for proteins such as ribosomal L9 and caldesmon [25,26]. The SAH domain of L9 is quite short and the sequence is not as rich in E, K and R residues as the SAH domain of caldesmon or myosin 6 ( Fig.…”

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confidence: 99%

Myosin tails and single α-helical domains

Batchelor

Wolny

Dougan

et al. 2015

Biochemical Society Transactions

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The human genome contains 39 myosin genes, divided up into 12 different classes. The structure, cellular function, and biochemical properties of many of these isoforms remains poorly characterized and there is still some controversy as to whether some myosin isoforms are monomers or dimers. Myosin isoforms 6 and 10 contain a stable single α-helical (SAH) domain, situated just after the canonical lever. The SAH domain is stiff enough to be able to lengthen the lever allowing the myosin to take a larger step. In addition, atomic force microscopy and atomistic simulations show that SAH domains unfold at relatively low forces and have a high propensity to refold. These properties are likely to be important for protein function, enabling motors to carry cargo in dense actin networks, and other proteins to remain attached to binding partners in the crowded cell.

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An exceptionally stable helix from the ribosomal protein L9: implications for protein folding and stability

Cited by 49 publications

References 51 publications

The Predicted Coiled-coil Domain of Myosin 10 Forms a Novel Elongated Domain That Lengthens the Head

The Predicted Coiled-coil Domain of Myosin 10 Forms a Novel Elongated Domain That Lengthens the Head

Amide proton exchange measurements as a probe of the stability and dynamics of the n‐terminal domain of the ribosomal protein L9: Comparison with the intact protein

Myosin tails and single α-helical domains

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