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SEC-SAXS experiments provided a direct
measurement of the global dimensions of the
A1-LCD (Fig. 1C). Guinier transformation of the
SAXS data yields an ensemble-averagedRg
value of 26.1 ± 1.1 Å (fig. S3). We extracted an
apparent scaling exponent ofnapp=0.45from
the experimental scattering data by fitting
to an empirically derived molecular form
factor (MFF) ( 30 ) (fig. S3). Results from all-
atom simulations agree with data from SEC-
SAXS experiments (Fig. 1C and fig. S4). The
distribution ofRgvalues obtained for the A1-
LCD from all-atom simulations (movie S1) is

biased toward compact conformations when

compared with those of self-avoiding random

walks and polymers at their theta temper-

atures (Fig. 1D). These analyses lead us to con-

clude that the A1-LCD adopts ensembles that

lie in the crossover regime between poor and

theta solvents.

We used NMR spectroscopy to uncover the

putative stickers that determine the features

of the conformational ensembles of the A1-

LCD. NMR transverse relaxation rates (R 2 )are

sensitive to internal motions slower than the

rotational correlation time and can be used to

identify regions of restricted motion. Fitting

the experimentally measuredR 2 rates as a func-

tion of sequence position within the A1-LCD

to a simple Gaussian chain model requires the

use of unrealistic values for the persistence

length (Fig. 1E). This suggests that groups of

residues have enhanced relaxation. Allowing

groups of enhancedR 2 rates along the se-

quence gave good agreement with the exper-

imental data using values for the persistence

length that are in line with those reported

previously for denatured proteins ( 31 ). Fixing

the group centers at aromatic residues results

Martinet al.,Science 367 , 694–699 (2020) 7 February 2020 2of6

Fig. 1. Aromatic residues are the stickers in the

PLD derived from hnRNPA1.(A) The sequence of the

PLD or LCD from hnRNPA1 (A1-LCD); aromatic residues

are indicated in orange. (B)^1 H-^15 NHSQCspectrum

recorded at 800 MHz and 25°C in pH 6 MES

buffer. For assignments see fig. S2A.w,chemical

shift; ppm, parts per million. (C)SEC-SAXS

data for A1-LCD. Calculatedscattering profiles from

simulated ensembles are overlaid in red.I(q)/I 0 ,

scattering intensity normalized by zero-angle scattering;

q, the momentum transfer vector, which is related to

scattering angle. (D)Rgdistribution from all-atom

simulations of A1-LCD (black) versus Gaussian chain

(violet) and self-avoiding random walk (SARW, green)

reference states. P(Rg), probability density distribution of

Rg.(E)^15 N amide transverse (R 2 ) relaxation rates

recorded at 800 MHz and 25°C. Overlaid are fits to the

data assuming a pure Gaussian-like profile (blue dashed

line) or multiple regions of enhanced relaxation centered

at aromatic residues (black dashed line) and the

underlying Gaussian-like profile from this fit (gray dashed

line) with a persistence length of 7.8 amino acid residues.

The yellow circles indicate the positions of the aromatic

residues. Gray bars indicate positions for which data were

not analyzed owing to unresolvable overlap in 2D spectra.

Monte Carlo sampling of the location of group centers

shows a clear positive correlation between the quality of

fit and positions of aromatic amino acids within the

sequence (fig. S5A). (F)^13 C-^1 H planes from the

aromatic-edited 3D NOESY (red) recorded at 800 MHz

and 25°C. Planes correspond to the Phe^1 Hd/e/z(left)

and Tyr^1 He(right) frequencies and their corresponding

diagonal signals are indicated by arrows. In both planes,

red boxes show signals at Tyr^1 Hd/^1 He(left) and Phe

(right), which exist in this plane only because of NOE

transfer. The^1 H,^13 C-HSQC aromatic region is shown

superimposed in blue. [For NOEs in a A1-LCD variant with

uniformly spaced aromatic residues (AroPerfect), see fig.

S5, E and F.] (G) Contact order from simulations. The

dashed lines and yellow circles indicate the positions

of all aromatic amino acids. (H) Normalized intensity

of Tyr-Phe NOEs as a function of temperature. The

Tyr^1 He–Phe NOE is displayed normalized to the

Tyr^1 Hd–^1 HeNOE of fixed distance at 5°, 15°, and 25°C. The

dashed line is a power-law fit. Single-letter abbreviations

for the amino acid residues are as follows: A, Ala; D,

Asp; F, Phe; G, Gly; K, Lys; M, Met; N, Asn; P, Pro; Q,

Gln; R, Arg; S, Ser; and Y, Tyr.

A

B

8.6 8.4 8.2 8.0 7.8 7.6

125

120

115

110

2 -

(^1) H (ppm)
1

• 
  

15

N (ppm)

10 -2

C 100

I(q)/I

0

q (Å-1)

0.01 0.1

ABSINTH simulation

form factor

Experimental uncertainty

A1-LCD

D

20 40 60

Rg (Å)

0

0.1

P(R

)g

A1-LCD

Gaussian chain

(at T)

SARW

E F

residue

contact order (10

–3

)

2

4

6

20 40 60 80 100 120

G

H

Tyr/Phe

temperature (oC)

51525

0.02

0.06

0.1

residue

20 40 60 80 100 120

R

(s 2

-1)

4

1

2

3

Single component Gaussian fit

Gaussian component, cluster fit

Multi-component cluster fit

7.2 7.0 6.8 6.6 6.4

2 -

(^1) H (ppm)
130
125
120
115
1

• 13

C (ppm)

7.0 6.8 6.6 6.4

Y

Y

F

Phe plane Tyr ε plane

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