Thermodynamics

The thermodynamic parameters at 15°C are summarized in Table 1. The number of sFcyRIIIa bound to an IgG1 molecule (N) was shown to be one. dFu-IgG1 had a 20 to 30 times higher binding constant for sFcyRIIIa than STD-IgG1, while AAA-IgG1 had a 3 to 4 times higher binding constant for sFcyRIIIa than STD-IgG1. IgG1-sFcyRIIIa binding was driven by favorable binding enthalpy (AH) but opposed by unfavorable binding entropy change (AS). IgG1-defucosylation improves its affinity for FcyRIIIa by enhancing the favorable AH, which contrasts markedly with the entropy-driven improvement of affinity by the amino acid substitution.

Table 1. Thermodynamic parameters for IgGl-sFcyRIIIIa(V) interaction

IgG1

N

Ka

AG

AH

-TAS

ACp

(x 106)

(kcal/mol)

(kcal/mol)

(kcal/mol/K)

(kcal/mol/K)

STD

1.1

1.87±0.07

-8.27±0.03

-20.1±0.5

11.8±0.4

-0.30±0.04

AAA

1.0

6.74±0.73

-9.00±0.06

-17.4±0.3

8.4±0.3

-0.33±0.04

dFu

1.0

58.3 ±11.3

-10.22±0.12

-26.1±0.7

15.8±0.8

-0.28±0.07

Favorable change in binding enthalpy generally indicates the increase in the non-covalent interactions, such as hydrogen bonds, van der Waals contacts, and salt bridges. Hence, the favorable change in AH brought about by the defucosylation suggests the increase in those non-covalent interactions in the complex.

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