pH Dependence of structural stability of interleukin-2 and granulocyte colony-stimulating factor.


Abstract

After a cytokine binds to its receptor on the cell surface (pH approximately 7), the complex is internalized into acidic endosomal compartments (pH approximately 5-6), where partially unfolded intermediates can form. The nature of these structural transitions was studied for wild-type interleukin-2 (IL-2) and wild-type granulocyte colony-stimulating factor (G-CSF). A noncoincidence of denaturation transitions in the secondary and tertiary structure of IL-2 and tertiary structural perturbations in G-CSF suggest the presence of an intermediate state for each, a common feature of this structural family of four-helical bundle proteins. Unexpectedly, both IL-2 and G-CSF display monotonic increases in stability as the pH is decreased from 7 to 4. We hypothesize that such cytokines with cell-based clearance mechanisms in vivo may have evolved to help stabilize endosomal complexes for sorting to lysosomal degradation. We show that mutants of both IL-2 and G-CSF have differential stabilities to their wild-type counterparts as a function of pH, and that these differences may explain the differences in ligand trafficking and depletion. Further understanding of the structural changes accompanying unfolding may help guide cytokine design with respect to ligand binding, endocytic trafficking, and, consequently, therapeutic efficacy. Study holds ProTherm entries: 16246, 16247, 16248, 16249, 16250, 16251, 16252, 16253, 16254, 16255, 16256, 16257, 16258, 16259, 16260, 16261, 16262, 16263, 16264, 16265 Extra Details: protein folding; protein stability; equilibrium denaturation; folding intermediates;,endocytic trafficking; IL-2; G-CSF

Submission Details

ID: DAu82sJH4

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:47 p.m.

Version: 1

Publication Details
Ricci MS;Sarkar CA;Fallon EM;Lauffenburger DA;Brems DN,Protein Sci. (2003) pH Dependence of structural stability of interleukin-2 and granulocyte colony-stimulating factor. PMID:12717025
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1IRL 1995-12-07 THE SOLUTION STRUCTURE OF THE F42A MUTANT OF HUMAN INTERLEUKIN 2
1GNC 1994-07-31 STRUCTURE AND DYNAMICS OF THE HUMAN GRANULOCYTE COLONY-STIMULATING FACTOR DETERMINED BY NMR SPECTROSCOPY. LOOP MOBILITY IN A FOUR-HELIX-BUNDLE PROTEIN
5GW9 2017-09-13 1.65 Crystal structure of C163, a backbone circularized G-CSF
5ZO6 2019-04-17 1.7 Crystal structure of C166, a backbone circularized G-CSF
4NEJ 2014-11-19 1.92 Small molecular fragment bound to crystal contact interface of Interleukin-2
4NEM 2014-11-19 1.93 Small molecular fragment bound to crystal contact interface of Interleukin-2
5LQB 2016-12-14 1.95 Complex structure of human IL2 mutant, Proleukin, with Fab fragment of NARA1 antibody
1M48 2002-07-31 1.95 Crystal Structure of Human IL-2 Complexed with (R)-N-[2-[1-(Aminoiminomethyl)-3-piperidinyl]-1-oxoethyl]-4-(phenylethynyl)-L-phenylalanine methyl ester
1M47 2002-07-31 1.99 Crystal Structure of Human Interleukin-2
6BFS 2018-09-12 2.0 The mechanism of GM-CSF inhibition by human GM-CSF auto-antibodies
1M49 2002-07-31 2.0 Crystal Structure of Human Interleukin-2 Complexed with SP-1985
5D70 2015-10-14 2.06 Crystal structure of MOR03929, a neutralizing anti-human GM-CSF antibody Fab fragment in complex with human GM-CSF
1M4B 2002-07-31 2.15 Crystal Structure of Human Interleukin-2 K43C Covalently Modified at C43 with 2-[2-(2-Cyclohexyl-2-guanidino-acetylamino)-acetylamino]-N-(3-mercapto-propyl)-propionamide
1M4A 2002-07-31 2.18 Crystal Structure of Human Interleukin-2 Y31C Covalently Modified at C31 with (1H-Indol-3-yl)-(2-mercapto-ethoxyimino)-acetic acid
1RHG 1994-01-31 2.2 THE STRUCTURE OF GRANULOCYTE-COLONY-STIMULATING FACTOR AND ITS RELATIONSHIP TO THOSE OF OTHER GROWTH FACTORS
1NBP 2002-12-18 2.2 Crystal Structure Of Human Interleukin-2 Y31C Covalently Modified At C31 With 3-Mercapto-1-(1,3,4,9-tetrahydro-B-carbolin-2-yl)-propan-1-one
5D71 2015-10-14 2.25 Crystal structure of MOR04302, a neutralizing anti-human GM-CSF antibody Fab fragment in complex with human GM-CSF
5M5E 2017-05-03 2.3 Crystal structure of a interleukin-2 variant in complex with interleukin-2 receptor
2B5I 2005-11-29 2.3 cytokine receptor complex
1M4C 2002-07-31 2.4 Crystal Structure of Human Interleukin-2
2GMF 1996-11-08 2.4 HUMAN GRANULOCYTE MACROPHAGE COLONY STIMULATING FACTOR
3INK 1993-10-31 2.5 UNRAVELING THE STRUCTURE OF INTERLEUKIN-2: REPLY
6BFQ 2018-09-12 2.6 The mechanism of GM-CSF inhibition by human GM-CSF auto-antibodies
1PW6 2004-01-13 2.6 Low Micromolar Small Molecule Inhibitor of IL-2
5D72 2015-10-14 2.6 Crystal structure of MOR04252, a neutralizing anti-human GM-CSF antibody Fab fragment in complex with human GM-CSF
4RS1 2015-11-11 2.68 Crystal structure of receptor-cytokine complex
1QVN 2005-04-05 2.7 Structure of SP4160 Bound to IL-2 V69A
1CSG 1993-10-31 2.7 Three-dimensional structure of recombinant human granulocyte-macrophage colony-stimulating factor
5UTZ 2018-07-11 2.75 Human IL-2/Fab complex
2D9Q 2006-02-07 2.8 Crystal Structure of the Human GCSF-Receptor Signaling Complex
1PY2 2004-01-13 2.8 Structure of a 60 nM Small Molecule Bound to a Hot Spot on IL-2
1CD9 2000-03-08 2.8 2:2 COMPLEX OF G-CSF WITH ITS RECEPTOR
1Z92 2005-06-07 2.8 structure of interleukin-2 with its alpha receptor
5C7X 2015-10-14 2.95 Crystal structure of MOR04357, a neutralizing anti-human GM-CSF antibody Fab fragment in complex with human GM-CSF
2ERJ 2006-02-21 3.0 Crystal structure of the heterotrimeric interleukin-2 receptor in complex with interleukin-2
3QB1 2012-04-11 3.1 Interleukin-2 mutant D10
4NKQ 2015-09-23 3.3 Structure of a Cytokine Receptor Complex
1PGR 2000-03-08 3.5 2:2 COMPLEX OF G-CSF WITH ITS RECEPTOR
3QAZ 2012-04-11 3.8 IL-2 mutant D10 ternary complex

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
98.3 Granulocyte colony-stimulating factor P09919 CSF3_HUMAN
91.0 Interleukin-2 Q8MKH2 IL2_SAISC
91.8 Interleukin-2 Q9XS38 IL2_PAPHA
91.8 Interleukin-2 Q7JFM4 IL2_AOTVO
91.8 Interleukin-2 Q7JFM3 IL2_AOTNI
91.8 Interleukin-2 Q7JFM5 IL2_AOTNA
91.8 Interleukin-2 Q7JFM2 IL2_AOTLE
94.8 Interleukin-2 Q29615 IL2_MACFA
95.5 Interleukin-2 P46649 IL2_CERAT
95.5 Interleukin-2 Q865Y1 IL2_PAPAN
96.3 Interleukin-2 P68290 IL2_MACNE
96.3 Interleukin-2 P68291 IL2_MACMU
99.2 Interleukin-2 P60569 IL2_HYLLA
99.2 Interleukin-2 P60568 IL2_HUMAN
94.4 Granulocyte-macrophage colony-stimulating factor Q0MUT8 CSF2_CHLAE
100.0 Granulocyte-macrophage colony-stimulating factor P04141 CSF2_HUMAN