The high- and low-affinity receptor binding sites of growth hormone are allosterically coupled

Walsh ST, Sylvester JE, Kossiakoff AA

Proc. Natl. Acad. Sci. U.S.A. 2004 Dec;101(49):17078-83

PMID: 15563602

Abstract

Growth hormone regulates its biological properties via a sequential hormone-induced receptor homodimerization mechanism. Using a mutagenesis-scanning analysis of 81 single and 32 pairwise double mutations, we show that the hormone’s two spatially distal receptor binding sites (Site1 and Site2) …

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The structural basis for biological signaling, regulation, and specificity in the growth hormone-prolactin system of hormones and receptors

Kossiakoff AA

Adv. Protein Chem. 2004;68:147-69

PMID: 15500861

Abstract

The pituitary hormones growth hormone (GH), prolactin (PRL) and placental lactogen (PL), are members of an extensive cytokine superfamily of hormones and receptors that share many of the same general structure-function relationships in expressing their biological activities. The biology of the pituitary hormones involves a very …

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Dissecting the binding energy epitope of a high-affinity variant of human growth hormone: cooperative and additive effects from combining mutations from independently selected phage display mutagenesis libraries

Bernat B, Sun M, Dwyer M, Feldkamp M, Kossiakoff AA

Biochemistry 2004 May;43(20):6076-84

PMID: 15147191

Abstract

Phage display mutagenesis is a widely used approach to engineering novel protein properties and is especially powerful in probing structure-function relationships in molecular recognition processes. The relative contributions of additive and cooperative binding forces and the influence of conformational …

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Site2 binding energetics of the regulatory step of growth hormone-induced receptor homodimerization

Walsh ST, Jevitts LM, Sylvester JE, Kossiakoff AA

Protein Sci. 2003 Sep;12(9):1960-70

PMID: 12930995

Abstract

Receptor signaling in the growth hormone (GH)-growth hormone receptor (GHR) system is controlled through a sequential two-step hormone-induced dimerization of two copies of the extracellular domain (ECD) of the receptor. The regulatory step of this process is the binding of …

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The functional binding epitope of a high affinity variant of human growth hormone mapped by shotgun alanine-scanning mutagenesis: insights into the mechanisms responsible for improved affinity

Pal G, Kossiakoff AA, Sidhu SS

J. Mol. Biol. 2003 Sep;332(1):195-204

PMID: 12946357

Abstract

A high-affinity variant of human growth hormone (hGH(v)) contains 15 mutations within site 1 and binds to the hGH receptor (hGHR) approximately 400-fold tighter than does wild-type (wt) hGH (hGH(wt)). We used shotgun scanning combinatorial mutagenesis to dissect the energetic contributions …

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The first semi-synthetic serine protease made by native chemical ligation

Pál G, Santamaria F, Kossiakoff AA, Lu W

Protein Expr. Purif. 2003 Jun;29(2):185-92

PMID: 12767808

Abstract

Selective incorporation of non-natural amino acid residues into proteins is a powerful approach to delineate structure-function relationships. Although many methodologies are available for chemistry-based protein engineering, more facile methods are needed to make this approach suitable for routine laboratory …

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Determination of the energetics governing the regulatory step in growth hormone-induced receptor homodimerization

Bernat B, Pal G, Sun M, Kossiakoff AA

Proc. Natl. Acad. Sci. U.S.A. 2003 Feb;100(3):952-7

PMID: 12552121

Abstract

Signaling in the human growth hormone (hGH)-human GH receptor system is initiated by a controlled sequential two-step hormone-induced dimerization of two hGH receptors via their extracellular domains (ECDs). Little is currently known about the energetics governing the …

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Structure of a phage display-derived variant of human growth hormone complexed to two copies of the extracellular domain of its receptor: evidence for strong structural coupling between receptor binding sites

Schiffer C, Ultsch M, Walsh S, Somers W, de Vos AM, Kossiakoff A

J. Mol. Biol. 2002 Feb;316(2):277-89

PMID: 11851338

Abstract

The structure of the ternary complex between the phage display- optimized, high-affinity Site 1 variant of human growth hormone (hGH) and two copies of the extracellular domain (ECD) of the hGH receptor (hGHR) has …

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High affinity RNase S-peptide variants obtained by phage display have a novel “hot-spot” of binding energy

Dwyer JJ, Dwyer MA, Kossiakoff AA

Biochemistry 2001 Nov;40(45):13491-500

PMID: 11695896

Abstract

Using phage display mutagenesis, high affinity variants of RNase S-peptide were produced that bind to RNase S-protein over 100-fold more tightly than the wild type S-peptide. The S-peptide: S-protein interface was further characterized using “biased” phage display libraries, where each targeted residue was …

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Biosynthetic phage display: a novel protein engineering tool combining chemical and genetic diversity

Dwyer MA, Lu W, Dwyer JJ, Kossiakoff AA

Chem. Biol. 2000 Apr;7(4):263-74

PMID: 10780926

Abstract

BACKGROUND: Molecular diversity in nature is developed through a combination of genetic and chemical elements. We have developed a method that permits selective manipulation of both these elements in one protein engineering tool. It combines the ability to introduce non-natural …

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