On Thursday it was announced that Bottas will be retained by the Silver Arrows for 2021 and, at a sweltering and deserted Silverstone, the Finn edged out Hamilton by just 0.136 seconds.
Red Bull's Max Verstappen was third ahead of Nico Hulkenberg, who is competing for Racing Point again following Sergio Perez's continued absence with coronavirus.
Hamilton heads into the second race in consecutive weeks at Silverstone with a 30-point championship lead following last weekend's British Grand Prix triumph.
Mercedes were on course to cruise to yet another one-two finish five days ago, before both of their drivers were subjected to late tyre failures.
Hamilton limped over the line to take a fortunate victory, but Bottas finished outside of the points.
Here, Mercedes continued their imperious form which has seen them win all four rounds so far, with Verstappen a distant seven tenths off Bottas' pace.
Hulkenberg was handed the keys to Perez's Racing Point on Friday morning – and, after he failed to start last weekend's grand prix following a car failure, he ended the running just behind Verstappen.
Ferrari's Charles Leclerc was the only other driver to finish within one second of Bottas – the Monegasque fifth in the order, 0.896 sec off the top.
The London-born Alexander Albon, whose seat at Red Bull is under scrutiny following a difficult start to the year, ended the running in sixth, 1.1 sec adrift. Four-time world champion Sebastian Vettel finished seventh, half-a-second off the pace of team-mate Leclerc.
Elsewhere, British driver Lando Norris, fourth in the standings ahead of Sunday's landmark grand prix, was 12th, two places ahead of McLaren team-mate Carlos Sainz. George Russell finished 15th for Williams.
The second 90-minute practice session gets under way at 3pm.