Table 2 Binding affinities and thermostability of the mutant clones selected by yeast display

From: SARS-CoV-2 variant prediction and antiviral drug design are enabled by RBD in vitro evolution

Clone

Library

Plasmida

Mutations

Tm (°C)

KD,YD (pM)c

KD (M)d

kon (M−1s−1)

koff (s−1)

Chi² (RU²)

RBD-WT

 

pJYDC1

Residues 336–528

53.9

1,700 ± 137 [8]

1.7 × 10−8

4.5 × 105

7.6 × 10−3

0.222

RBD-32

B3

pJYDC1

I358F, S477N, N501Y

ND

348 ± 35 [3]

ND

   

RBD-33

B3

pJYDC1

I358F, E484K, N501Y

ND

204 ± 88 [4]

ND

   

RBD-36

B3

pJYDC1

I358F, I468T, N481Y, N501Y

54.6

268 ± 35 [3]

1.0 × 10−8

2.1 × 105

2.2 × 10−3

0.119

RBD-48

B4

pJYDC3

I358F, E484K, Q498R, N501Y

54.8

55 ± 23 [4]

1.7 × 10−10

1.0 × 106

1.7 × 10−4

0.182

RBD-52

B5

pJYDC1

I358F, N460K, E484K, S494P, Q498R, N501Y, A520G

61.9

40 ± 16 [5]

2.4 × 10−10

3.8 × 105

9.0 × 10−5

0.0515

RBD-521

B5

pJYDC1

I358F, N460K, E484K, Q498R, N501Y

58.3

14 ± 3 [3]

1.7 × 10−10

9.0 × 105

1.6 × 10−4

0.0562

RBD-62

B6(FA)

pJYDC3

I358F, V445K, N460K, I468T, T470M, S477N, E484K, Q498R, N501Y

57.6

3.5 ± 1.8 [4]

1.6 × 10−11

4.4 × 106

6.8 × 10−5

0.42

RBD-614

B6(FA)

pJYDC3

I358F, V445K, N460K, E484K, S494P, Q498R, N501Y

58.5

21 ± 6.2 [3]

1.9 × 10−10

6.8 × 105

1.3 × 10−4

0.0488

RBD-71

 

pJYDC3

I358F, V367W, R408D, K417V, V445K, N460K, I468T, T470M, S477N, E484K, Q498R, N501Y

63.5

8.5 ± 2.5 [3]

1.6 × 10−10

1.2 × 106

2.0 × 10−4

0.104

  1. Underlined clones have the highest affinity among the clones of a given library, and their representative binding curves are shown in Fig. 2a. ND, not determined; RU, response units.
  2. apJYDC1 (plasmid 162458) uses the intrinsic eUnaG2 reporter; pJYDC3 (plasmid 162460) contains the DnbALFA reporter21.
  3. bMelting temperature (Tm) as measured by differential scanning fluorimeter Tycho NT.6 (NanoTemper Technologies).
  4. cKD values measured between yeast surface-exposed RBD variants and the monomeric extracellular portion of the ACE2 receptor (residues G17–Y613). The number of biological replicates is shown in square brackets.
  5. dMeasured by the Biacore S200 (GE Healthcare) using a series S sensor chip protein A (GE Healthcare) activated with Fc–ACE227 (Methods). The sensorgrams are shown in Fig. 2h (WT and RBD-62) and Extended Data Fig. 5. Values were calculated from global fitting of the data. kon and koff, association and dissociation rate constants, respectively; Chi2, Chi squared test value.
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