Structure-Activity Studies of Interleukin-2

Cohen, FE; Kosen, PA; Kuntz, I. D.; Lb, Epstein; Ciardelli, TL; Smith, K. A.

doi:10.1126/science.3489989

Cited by 115 publications

(33 citation statements)

References 28 publications

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“…Consequently, the results presented here describe the intrinsic tendencies of four individual helices to pack into the four-a-helix bundle in the absence of any additional constraints (2) that may arise from chain connectivity. There are three possible ways of connecting the helices in an antiparallel four-a-helix bundle, all of which have been observed in proteins (13), as well as several different arrangements of the successive helices in the bundle (34,35). The present conclusions apply to bundles with any kind of connectivity.…”

Section: Methodssupporting

confidence: 47%

Energetics of the structure of the four-alpha-helix bundle in proteins.

Chou

Maggiora

Némethy

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

124

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The main features of the four-a-helix bundle, one of the characteristic structural elements of many proteins, can be explained in terms of noncovalent interactions between the constituent helices. Conformational energy computations have been carried out on four types of four-a-helix bundles, each consisting of four CH3CO-(L-Ala)1O-NHCH3 polypeptide chains, with various combinations of parallel and antiparallel orientations of the helices. In the bundle with the most favorable energy, all pairs of neighboring helices are oriented antiparallel-i.e., in the orientation that is favored by electrostatic interactions between the helices. In this structure, the orientation angle between neighboring helix axes is -168°, within ± 70, in close agreement with the orientation angles observed in proteins and with the value that we computed earlier for the most favorable packing of pairs of interacting a-helices. This orientation corresponds to a left-handed twisting of the helical bundle. The preferred handedness of this twisting arises as a result of favorable nonbonded interactions between the a-helices.

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Section: Methodssupporting

confidence: 47%

Energetics of the structure of the four-alpha-helix bundle in proteins.

Chou

Maggiora

Némethy

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

124

View full text Add to dashboard Cite

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“…As noted earlier, Weber and Salemme (8) proposed that all four-a-helix bundles were right-handed. Thus, a right-handed structure was selected for interleukin 2 [a left-handed four-a-helix bundle (31)], even though the combinatoric algorithms produced left-handed and right-handed structures (44). This report demonstrates that left-handed and right-handed structures occur with approximately equal frequency.…”

Section: Resultsmentioning

confidence: 90%

Topological distribution of four-alpha-helix bundles.

Presnell

Cohen

1989

Proc. Natl. Acad. Sci. U.S.A.

182

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The four-a-helix bundle, a common structural motif in globular proteins, provides an excellent forum for the examination of predictive constraints for protein backbone topology. An exhaustive examination of the Brookhaven Crystallographic Protein Data Bank and other literature sources has lead to the discovery of 20 putative four-a-helix bundles. Application of an analytical method that examines the difference between solvent-accessible surface areas in packed and partially unpacked bundles reduced the number of structures to 16. Angular requirements further reduced the list of bundles to 13. In 12 of these bundles, all pairs of neighboring helices were oriented in an anti-parallel fashion. This distribution is in accordance with structure types expected if the helix macro dipole effect makes a substantial contribution to the stability of the native structure. The characterizations and classifications made in this study prompt a reevaluation of constraints used in structure prediction efforts.Specification of the code that translates primary to tertiary structure remains unresolved. It has proven difficult to predict which features in an amino acid sequence will provide the basis for the three-dimensional structure or function of a given protein. However, proteins with only 10% identity at comparable positions in a polypeptide can have notably similar structures (1). Indeed, an individual tertiary structure will often fall into one of a limited number of structural classes (2). Tertiary-structure prediction systems incorporating combinatorial methods (3-5) and template methods (6, 7) target known structural classes in an attempt to reduce the number of structures created and examined. Therefore, to effectively predict tertiary structures, we must determine as many constraints describing the individual structural classes as possible.Among the structural class containing those proteins constructed predominantly from a-helical structures, a highly recurrent-motif is the collection of (anti)parallel a-helices known as the four-a-helix bundle. Four-a-helix bundles are found in proteins covering a wide range of structure and function, but they display some common characteristics. Weber and Salemme (8) were among the first to examine and characterize four-a-helix bundles as a class of supersecondary structure. Initially, their work suggested a common, right-handed, connective topology for all four-a-helix bundles. This characterization, derived from a limited data base of bundle topologies, proved too restrictive to be correct. The "handedness" constraint obscured further attempts to characterize the topology of previously known and newly discovered structures (9, 10). Incorporation of all the presently known four-a-helix bundle structures requires a different categorization scheme.The purpose of this study is to describe and present a thorough examination of the literature for four-a-helix bundles using generalized determination criteria. These criteria suggest a set of bundle structures beyond those initially...

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“…According to fluorescence quenching experiments this Trp is buried [31,32]. During denaturation in concentrated guanidinium chloride solution the maximum was shifted rapidly, i.e.…”

Section: Stability Of Muteins In Guunidinium Chloridementioning

confidence: 99%

Mutant proteins of human interleukin 2

Weigel

Meyer

Sebald

1989

European Journal of Biochemistry

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Muteins, i.e. proteins altered by mutation of their genes, of interleukin 2 (112) were generated by oligonucleotide-directed mutagenesis in vitro. All acidic and basic residues conserved between man and mouse were exchanged as well as four lipophilic residues contained within four hydrophobic segments of the protein.The muteins were produced in Escherichia coli and submitted to a renaturation and purification protocol, before bioactivity and receptor binding of each of them was determined.All muteins besides two (K44/T125 and QIlO/T125) could be renatured and purified. One mutein (K94/T12.5) exhibited a more than tenfold-improved renaturation yield.One amino exchange (Asp-20 to Asn) resulted in an about 20-fold reduction in proliferative activity and highaffinity receptor binding. The binding to the low-affinity 112-binding protein (Tac antigen) was unimpaired.A second exchange (Arg-38 to Gln) had no effect on proliferative activity. The binding to both the high-and the low-affinity receptor, however, was reduced about 20-fold.Preliminary trials on the stability of these muteins by guanidinium hydrochloride denaturation studies detected no differences between wild-type interleukin 2 and muteins.It is suggested that Asp-20 forms part of the binding site for the large receptor subunit whereas Arg-38 is involved in the contact site to the small subunit.The transient synthesis of interleukin 2 (112) and its receptor by activated T lymphocytes apparently represents an essential step in the elaboration of the cellular immune response (for recent reviews see [I -31). The binding of 112 to a highaffinity receptor (& = 10 pM) provides the growth-signal leading to the clonal expansion of antigen-specific T cells.Interestingly, the high-affinity receptor has been established to be assembled from a low-affinity (Kd z 10 nM) 112- large extent preliminary, however, since the proteins were not renatured and purified before measuring proliferative or receptor-binding activity. Thus, effects of a certain mutation on the folding or on the receptor-binding sites could not be clearly differentiated.During the studies presented in this paper a series of charged residue positions and of hydrophobic positions were exchanged which cover more or less the whole surface of 112. The muteins were all submitted to a renaturation and purification protocol before analysing biological activities. Two muteins appeared which define positions involved in high-or in low-affinity receptor binding. Surprisingly, one mutein was obtained exhibiting a large increase in yield during renaturation. METHODS Construction of mutant strainsA cDNA coding for the mature part of human I12 [8] plus an initiator methionine [I91 was submitted to oligonucleotidedirected mutagenesis by the gapped duplex DNA approach [20, 211. The synthetic oligonucleotides comprised 6-16 residues before and after the nucleotide(s) to be exchanged, and they were provided by Dr H. Blocker (GBF, Braunschweig) or synthesised on a DNA synthesiser (Applied Biosystems, model 380). Mutation...

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Structure-Activity Studies of Interleukin-2

Cited by 115 publications

References 28 publications

Energetics of the structure of the four-alpha-helix bundle in proteins.

Energetics of the structure of the four-alpha-helix bundle in proteins.

Topological distribution of four-alpha-helix bundles.

Mutant proteins of human interleukin 2

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